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Sample records for capillary gel electrophoresis

  1. Capillary gel electrophoresis for rapid, high resolution DNA sequencing.

    OpenAIRE

    Swerdlow, H; Gesteland, R

    1990-01-01

    Capillary gel electrophoresis has been demonstrated for the separation and detection of DNA sequencing samples. Enzymatic dideoxy nucleotide chain termination was employed, using fluorescently tagged oligonucleotide primers and laser based on-column detection (limit of detection is 6,000 molecules per peak). Capillary gel separations were shown to be three times faster, with better resolution (2.4 x), and higher separation efficiency (5.4 x) than a conventional automated slab gel DNA sequenci...

  2. 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

  3. 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.

  4. Performance comparison of capillary and agarose gel electrophoresis for the identification and characterization of monoclonal immunoglobulins.

    Science.gov (United States)

    McCudden, Christopher R; Mathews, Stephanie P; Hainsworth, Shirley A; Chapman, John F; Hammett-Stabler, Catherine A; Willis, Monte S; Grenache, David G

    2008-03-01

    The objective of this study was to compare gel- and capillary-based serum protein electrophoresis methods to identify and characterize monoclonal immunoglobulins (M proteins). Five reviewers interpreted 149 consecutively ordered serum specimens following agarose gel electrophoresis (AGE), capillary electrophoresis (CE), immunofixation electrophoresis (IFE), and subtraction immunotyping (IT). As a screening test for detecting M proteins, AGE and CE displayed similar sensitivity (91% and 92%, respectively). CE was less specific (74%) than AGE (81%). An analysis of interinterpreter agreement revealed that interpretations were more consistent using gel-based methods than capillary-based methods, with 80% of the gel interpretations being in complete (5/5) agreement compared with 67% of the capillary interpretations. After implementing the capillary-based methods, the number of tests per reportable result increased (from 1.58 to 1.73). CE is an analytically suitable alternative to AGE, but laboratories implementing it will need to continue IFE testing to characterize all M proteins detected by CE.

  5. 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.

  6. Gel versus capillary electrophoresis genotyping for categorizing treatment outcomes in two anti-malarial trials in Uganda

    OpenAIRE

    Hubbard Alan E; Dorsey Grant; Gupta Vinay; Rosenthal Philip J; Greenhouse Bryan

    2010-01-01

    Abstract Background Molecular genotyping is performed in anti-malarial trials to determine whether recurrent parasitaemia after therapy represents a recrudescence (treatment failure) or new infection. The use of capillary instead of agarose gel electrophoresis for genotyping offers technical advantages, but it is unclear whether capillary electrophoresis will result in improved classification of anti-malarial treatment outcomes. Methods Samples were genotyped using both gel and capillary elec...

  7. Gel versus capillary electrophoresis genotyping for categorizing treatment outcomes in two anti-malarial trials in Uganda

    Directory of Open Access Journals (Sweden)

    Hubbard Alan E

    2010-01-01

    Full Text Available Abstract Background Molecular genotyping is performed in anti-malarial trials to determine whether recurrent parasitaemia after therapy represents a recrudescence (treatment failure or new infection. The use of capillary instead of agarose gel electrophoresis for genotyping offers technical advantages, but it is unclear whether capillary electrophoresis will result in improved classification of anti-malarial treatment outcomes. Methods Samples were genotyped using both gel and capillary electrophoresis from randomized trials of artemether-lumefantrine (AL vs. dihydroartemisinin-piperaquine (DP performed in two areas of Uganda: Kanungu, where transmission is moderate, and Apac, where transmission is very high. Both gel and capillary methods evaluated polymorphic regions of the merozoite surface protein 1 and 2 and glutamine rich protein genes. Results Capillary electrophoresis detected more alleles and provided higher discriminatory power than agarose gel electrophoresis at both study sites. There was only moderate agreement between classification of outcomes with the two methods in Kanungu (kappa = 0.66 and poor agreement in Apac (kappa = 0.24. Overall efficacy results were similar when using gel vs. capillary methods in Kanungu (42-day risk of treatment failure for AL: 6.9% vs. 5.5%, p = 0.4; DP 2.4% vs. 2.9%, p = 0.5. However, the measured risk of recrudescence was significantly higher when using gel vs. capillary electrophoresis in Apac (risk of treatment failure for AL: 17.0% vs. 10.7%, p = 0.02; DP: 8.5% vs. 3.4%, p = 0.03. Risk differences between AL and DP were not significantly different whether gel or capillary methods were used. Conclusions Genotyping with gel electrophoresis overestimates the risk of recrudescence in anti-malarial trials performed in areas of high transmission intensity. Capillary electrophoresis provides more accurate outcomes for such trials and should be performed when possible. In areas of moderate transmission

  8. Gel versus capillary electrophoresis genotyping for categorizing treatment outcomes in two anti-malarial trials in Uganda.

    Science.gov (United States)

    Gupta, Vinay; Dorsey, Grant; Hubbard, Alan E; Rosenthal, Philip J; Greenhouse, Bryan

    2010-01-15

    Molecular genotyping is performed in anti-malarial trials to determine whether recurrent parasitaemia after therapy represents a recrudescence (treatment failure) or new infection. The use of capillary instead of agarose gel electrophoresis for genotyping offers technical advantages, but it is unclear whether capillary electrophoresis will result in improved classification of anti-malarial treatment outcomes. Samples were genotyped using both gel and capillary electrophoresis from randomized trials of artemether-lumefantrine (AL) vs. dihydroartemisinin-piperaquine (DP) performed in two areas of Uganda: Kanungu, where transmission is moderate, and Apac, where transmission is very high. Both gel and capillary methods evaluated polymorphic regions of the merozoite surface protein 1 and 2 and glutamine rich protein genes. Capillary electrophoresis detected more alleles and provided higher discriminatory power than agarose gel electrophoresis at both study sites. There was only moderate agreement between classification of outcomes with the two methods in Kanungu (kappa = 0.66) and poor agreement in Apac (kappa = 0.24). Overall efficacy results were similar when using gel vs. capillary methods in Kanungu (42-day risk of treatment failure for AL: 6.9% vs. 5.5%, p = 0.4; DP 2.4% vs. 2.9%, p = 0.5). However, the measured risk of recrudescence was significantly higher when using gel vs. capillary electrophoresis in Apac (risk of treatment failure for AL: 17.0% vs. 10.7%, p = 0.02; DP: 8.5% vs. 3.4%, p = 0.03). Risk differences between AL and DP were not significantly different whether gel or capillary methods were used. Genotyping with gel electrophoresis overestimates the risk of recrudescence in anti-malarial trials performed in areas of high transmission intensity. Capillary electrophoresis provides more accurate outcomes for such trials and should be performed when possible. In areas of moderate transmission, gel electrophoresis appears adequate to estimate comparative

  9. Staining Method for Protein Analysis by Capillary Gel Electrophoresis

    Science.gov (United States)

    Wu, Shuqing; Lu, Joann J; Wang, Shili; Peck, Kristy L.; Li, Guigen; Liu, Shaorong

    2009-01-01

    A novel staining method and the associated fluorescent dye were developed for protein analysis by capillary SDS-PAGE. The method strategy is to synthesize a pseudo-SDS dye and use it to replace some of the SDS in SDS–protein complexes so that the protein can be fluorescently detected. The pseudo-SDS dye consists of a long, straight alkyl chain connected to a negative charged fluorescent head and binds to proteins just as SDS. The number of dye molecules incorporated with a protein depends on the dye concentration relative to SDS in the sample solution, since SDS and dye bind to proteins competitively. In this work, we synthesized a series of pseudo-SDS dyes, and tested their performances for capillary SDS-PAGE. FT-16 (a fluorescein molecule linked with a hexadodecyl group) seemed to be the best among all the dyes tested. Although the numbers of dye molecules bound to proteins (and the fluorescence signals from these protein complexes) were maximized in the absence of SDS, high-quality separations were obtained when co-complexes of SDS–protein–dye were formed. The migration time correlates well with protein size even after some of the SDS in the SDS–protein complexes was replaced by the pseudo-SDS dye. Under optimized experimental conditions and using a laser-induced fluorescence detector, limits of detection of as low as 0.13 ng/mL (bovine serum albumin) and dynamic ranges over 5 orders of magnitude in which fluorescence response is proportional to the square root of analyte concentration were obtained. The method and dye were also tested for separations of real-world samples from E. coli. PMID:17874848

  10. Sodium dodecyl sulfate-capillary gel electrophoresis of polyethylene glycolylated interferon alpha.

    Science.gov (United States)

    Na, Dong H; Park, Eun J; Youn, Yu S; Moon, Byung W; Jo, Yeong W; Lee, Sung H; Kim, Won-Bae; Sohn, Yeowon; Lee, Kang C

    2004-02-01

    Sodium dodecyl sulfate-capillary gel electrophoresis (SDS-CGE) using a hydrophilic replaceable polymer network matrix was applied to characterize the polyethylene glycol(PEG)ylated interferon alpha (PEG-IFN). The SDS-CGE method resulted in a clearer resolution in both the PEG-IFN species and the native IFN species. The distribution profile of PEGylation determined by SDS-CGE was consistent with that obtained by SDS-polyacrylamide gel electrophoresis (PAGE) with Coomassie blue or barium iodide staining. The result was also compared using matrix-assisted laser desorption/ionization-time of flight-mass spectrometry. SDS-CGE was also useful for monitoring the PEGylation reaction to optimize the reaction conditions, such as reaction molar ratio. This study shows the potential of SDS-CGE as a new method for characterizing the PEGylated proteins with advantages of speed, minimal sample consumption and high resolution.

  11. Determination of molecular weight of silk fibroin by non-gel sieving capillary electrophoresis.

    Science.gov (United States)

    Wei, Wei; Zhang, Yaopeng; Shao, Huili; Hu, Xuechao

    2010-01-01

    A simple non-gel sieving capillary electrophoresis (NGSCE) method was established to determine the MW of silk fibroin using CE. The background electrolyte with a pH of 8.8 was based on three components: polyethylene glycol, tris(hydroxymethyl)aminomethane, and sodium dodecyl sulfate (SDS). NGSCE showed a good linear relationship with satisfactory reproducibility between the migration time and the MW of standard proteins. It was found that the regenerated silk fibroin had an MW around 83 kDa with a wide MW distribution (MWD). This absolute value is lower than the result obtained from SDS-polyacrylamide gel electrophoresis due to the different principles of the methods, but their similar MWD shapes indicated that NGSCE could be a feasible, highly sensitive, rapid method for determination of the MW of silk fibroin.

  12. Western Blotting using Capillary Electrophoresis

    OpenAIRE

    Anderson, Gwendolyn J.; Cipolla, Cynthia; Kennedy, Robert T.

    2011-01-01

    A microscale Western blotting system based on separating sodium-dodecyl sulfate protein complexes by capillary gel electrophoresis followed by deposition onto a blotting membrane for immunoassay is described. In the system, the separation capillary is grounded through a sheath capillary to a mobile X-Y translation stage which moves a blotting membrane past the capillary outlet for protein deposition. The blotting membrane is moistened with a methanol and buffer mixture to facilitate protein a...

  13. Combining ligation reaction and capillary gel electrophoresis to obtain reliable long DNA probes.

    Science.gov (United States)

    García-Cañas, Virginia; Mondello, Monica; Cifuentes, Alejandro

    2011-05-01

    New DNA amplification methods are continuously developed for sensitive detection and quantification of specific DNA target sequences for, e.g. clinical, environmental or food applications. These new applications often require the use of long DNA oligonucleotides as probes for target sequences hybridization. Depending on the molecular technique, the length of DNA probes ranges from 40 to 450 nucleotides, solid-phase chemical synthesis being the strategy generally used for their production. However, the fidelity of chemical synthesis of DNA decreases for larger DNA probes. Defects in the oligonucleotide sequence result in the loss of hybridization efficiency, affecting the sensitivity and selectivity of the amplification method. In this work, an enzymatic procedure has been developed as an alternative to solid-phase chemical synthesis for the production of long oligonucleotides. The enzymatic procedure for probe production was based on ligation of short DNA sequences. Long DNA probes were obtained from smaller oligonucleotides together with a short sequence that acts as bridge stabilizing the molecular complex for DNA ligation. The ligation reactions were monitored by capillary gel electrophoresis with laser-induced fluorescence detection (CGE-LIF) using a bare fused-silica capillary. The capillary gel electrophoresis-LIF method demonstrated to be very useful and informative for the characterization of the ligation reaction, providing important information about the nature of some impurities, as well as for the fine optimization of the ligation conditions (i.e. ligation cycles, oligonucleotide and enzyme concentration). As a result, the yield and quality of the ligation product were highly improved. The in-lab prepared DNA probes were used in a novel multiplex ligation-dependent genome amplification (MLGA) method for the detection of genetically modified maize in samples. The great possibilities of the whole approach were demonstrated by the specific and sensitive

  14. Tunable separations based on a molecular size effect for biomolecules by poly(ethylene glycol) gel-based capillary electrophoresis.

    Science.gov (United States)

    Kubo, Takuya; Nishimura, Naoki; Furuta, Hayato; Kubota, Kei; Naito, Toyohiro; Otsuka, Koji

    2017-11-10

    We report novel capillary gel electrophoresis (CGE) with poly(ethylene glycol) (PEG)-based hydrogels for the effective separations of biomolecules containing sugars and DNAs based on a molecular size effect. The gel capillaries were prepared in a fused silica capillary modified with 3-(trimethoxysilyl)propylmethacrylate using a variety of the PEG-based hydrogels. After the fundamental evaluations in CGE regarding the separation based on the molecular size effect depending on the crosslinking density, the optimized capillary provided the efficient separation of glucose ladder (G1 to G20). In addition, another capillary showed the successful separation of DNA ladder in the range of 10-1100 base pair, which is superior to an authentic acrylamide-based gel capillary. For both glucose and DNA ladders, the separation ranges against the molecular size were simply controllable by alteration of the concentration and/or units of ethylene oxide in the PEG-based crosslinker. Finally, we demonstrated the separations of real samples, which included sugars carved out from monoclonal antibodies, mAbs, and then the efficient separations based on the molecular size effect were achieved. Copyright © 2017 Elsevier B.V. All rights reserved.

  15. Adapting capillary gel electrophoresis as a sensitive, high-throughput method to accelerate characterization of nucleic acid metabolic enzymes.

    Science.gov (United States)

    Greenough, Lucia; Schermerhorn, Kelly M; Mazzola, Laurie; Bybee, Joanna; Rivizzigno, Danielle; Cantin, Elizabeth; Slatko, Barton E; Gardner, Andrew F

    2016-01-29

    Detailed biochemical characterization of nucleic acid enzymes is fundamental to understanding nucleic acid metabolism, genome replication and repair. We report the development of a rapid, high-throughput fluorescence capillary gel electrophoresis method as an alternative to traditional polyacrylamide gel electrophoresis to characterize nucleic acid metabolic enzymes. The principles of assay design described here can be applied to nearly any enzyme system that acts on a fluorescently labeled oligonucleotide substrate. Herein, we describe several assays using this core capillary gel electrophoresis methodology to accelerate study of nucleic acid enzymes. First, assays were designed to examine DNA polymerase activities including nucleotide incorporation kinetics, strand displacement synthesis and 3'-5' exonuclease activity. Next, DNA repair activities of DNA ligase, flap endonuclease and RNase H2 were monitored. In addition, a multicolor assay that uses four different fluorescently labeled substrates in a single reaction was implemented to characterize GAN nuclease specificity. Finally, a dual-color fluorescence assay to monitor coupled enzyme reactions during Okazaki fragment maturation is described. These assays serve as a template to guide further technical development for enzyme characterization or nucleoside and non-nucleoside inhibitor screening in a high-throughput manner. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  16. Separation of oligopeptides, nucleobases, nucleosides and nucleotides using capillary electrophoresis/electrochromatography with sol–gel modified inner capillary wall

    Czech Academy of Sciences Publication Activity Database

    Svobodová, Jana; Kofroňová, Olga; Benada, Oldřich; Král, V.; Mikšík, Ivan

    2017-01-01

    Roč. 1517, Sep 29 (2017), s. 185-194 ISSN 0021-9673 R&D Projects: GA ČR(CZ) GA15-01948S; GA MŠk(CZ) LO1509 Institutional support: RVO:67985823 ; RVO:61388971 Keywords : capillary electrochromatography (CEC) * open-tubular capillary electrochromatography (OT-CEC) * nucleo-compounds * oligopeptides * sol–gel methods * Porphyrin * scanning electron microscopy (SEM) Subject RIV: CB - Analytical Chemistry, Separation; CE - Biochemistry (MBU-M) OBOR OECD: Analytical chemistry; Biochemistry and molecular biology (MBU-M) Impact factor: 3.981, year: 2016

  17. Coupling Sodium Dodecyl Sulfate–Capillary Polyacrylamide Gel Electrophoresis with MALDI-TOF-MS via a PTFE Membrane

    Science.gov (United States)

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

    2011-01-01

    Sodium dodecyl sulfate (SDS)–polyacrylamide gel electrophoresis (PAGE) is a fundamental analytical technique for proteomic research, and SDS–capillary gel electrophoresis (CGE) is its miniaturized version. Compared to conventional slab-gel electrophoresis, SDS-CGE has many advantages such as increased separation efficiency, reduced separation time and automated operation. SDS-CGE is not widely accepted in proteomic research primarily due to the difficulties in identifying the well-resolved proteins. MALDI–TOF–MS is an outstanding platform for protein identifications. Coupling the two would solve the problem but is extremely challenging because the MS detector has no access to the SDS-CGE resolved proteins and the SDS interferes with MS detection. In this work we introduce an approach to address these issues. We discover that poly(tetrafluoroethylene) (PTFE) membranes are excellent materials for collecting SDS-CGE separated proteins. We demonstrate that we can wash off the SDS bound to the collected proteins and identify these proteins on-membrane with MALDI-TOF-MS. We also show that we can immunoblot and Coomassie-stain the proteins collected on these membranes. PMID:21309548

  18. Western blotting using capillary electrophoresis.

    Science.gov (United States)

    Anderson, Gwendolyn J; M Cipolla, Cynthia; Kennedy, Robert T

    2011-02-15

    A microscale Western blotting system based on separating sodium-dodecyl sulfate protein complexes by capillary gel electrophoresis followed by deposition onto a blotting membrane for immunoassay is described. In the system, the separation capillary is grounded through a sheath capillary to a mobile X-Y translation stage which moves a blotting membrane past the capillary outlet for protein deposition. The blotting membrane is moistened with a methanol and buffer mixture to facilitate protein adsorption. Although discrete protein zones could be detected, bands were broadened by ∼1.7-fold by transfer to membrane. A complete Western blot for lysozyme was completed in about one hour with 50 pg mass detection limit from low microgram per milliliter samples. These results demonstrate substantial reduction in time requirements and improvement in mass sensitivity compared to conventional Western blots. Western blotting using capillary electrophoresis shows promise to analyze low volume samples with reduced reagents and time, while retaining the information content of a typical Western blot.

  19. Practical capillary electrophoresis

    CERN Document Server

    Weinberger, Robert

    2000-01-01

    In the 1980s, capillary electrophoresis (CE) joined high-performance liquid chromatography (HPLC) as the most powerful separation technique available to analytical chemists and biochemists. Published research using CE grew from 48 papers in the year of commercial introduction (1988) to 1200 in 1997. While only a dozen major pharmaceutical and biotech companies have reduced CE to routine practice, the applications market is showing real or potential growth in key areas, particularly in the DNA marketplace for genomic mapping and forensic identification. For drug development involving small molecules (including chiral separations), one CE instrument can replace 10 liquid chromatographs in terms of speed of analysis. CE also uses aqueous rather than organic solvents and is thus environmentally friendlier than HPLC. The second edition of Practical Capillary Electrophoresis has been extensively reorganized and rewritten to reflect modern usage in the field, with an emphasis on commercially available apparatus and ...

  20. 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

  1. Conducting polymer electrodes for gel electrophoresis.

    Directory of Open Access Journals (Sweden)

    Katarina Bengtsson

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

  2. Conducting polymer electrodes for gel electrophoresis.

    Science.gov (United States)

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

    2014-01-01

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

  3. Capillary gel electrophoresis-coupled aptamer enzymatic cleavage protection strategy for the simultaneous detection of multiple small analytes.

    Science.gov (United States)

    Perrier, Sandrine; Zhu, Zhenyu; Fiore, Emmanuelle; Ravelet, Corinne; Guieu, Valérie; Peyrin, Eric

    2014-05-06

    This novel, multi small-analyte sensing strategy is the result of combining the target-induced aptamer enzymatic protection approach with the CGE-LIF (capillary gel electrophoresis with laser-induced fluorescence) technique. The implemented assay principle is based on an analysis of the phosphodiesterase I (PDE I)-mediated size variation of a fluorescein-labeled aptamer (FApt), the enzyme catalyzing the removal of nucleotides from DNA in the 3' to 5' direction. In the absence of the target, the unfolded aptamer was enzymatically cleaved into short DNA fragments. Upon target binding, the DNA substrate was partially protected against enzymatic hydrolysis. The amount of bound aptamer remaining after the exonuclease reaction was proportional to the concentration of the target. The CGE technique, which was used to determine the separation of FApt species from DNA digested products, permitted the quantification of adenosine (A), ochratoxin A (O), and tyrosinamide (T) under the same optimized enzymatic conditions. This assay strategy was subsequently applied to the simultaneous detection of A, O, and T in a single capillary under buffered conditions using corresponding FApt probes of different lengths (23, 36, and 49 nucleotides, respectively). Additionally, the detection of these three small molecules was successfully achieved in a complex medium (diluted, heat-treated human serum) showing a good recovery. It is worth noting that the multiplexed analysis was accomplished for targets with different charge states by using aptamers possessing various structural features. This sensing platform constitutes a rationalized and reliable approach with an expanded potential for a high-throughput determination of small analytes in a single capillary.

  4. Biomedical applications of capillary electrophoresis

    International Nuclear Information System (INIS)

    Kartsova, L A; Bessonova, E A

    2015-01-01

    The review deals with modern analytical approaches used in capillary electrophoresis for solving medical and biological problems: search for biomarkers of various diseases and rapid diagnosis based on characteristic profiles of biologically active compounds by capillary electrophoresis with mass spectrometric detection; monitoring of the residual drugs in biological fluids for evaluating the efficiency of drug therapy; testing of the enantiomeric purity of pharmaceutical products; the use of novel materials as components of stationary and pseudo-stationary phases in capillary electrophoresis and capillary electrochromatography to increase the selectivity of separation of components of complex matrices; and identification of various on-line preconcentration techniques to reduce the detection limits of biologically active analytes. A topical trend in capillary electrophoresis required in clinical practice, viz., the design of microfluidic systems, is discussed. The bibliography includes 173 references

  5. DNA Sequencing by Capillary Electrophoresis

    Science.gov (United States)

    Karger, Barry L.; Guttman, Andras

    2009-01-01

    Sequencing of human and other genomes has been at the center of interest in the biomedical field over the past several decades and is now leading toward an era of personalized medicine. During this time, DNA sequencing methods have evolved from the labor intensive slab gel electrophoresis, through automated multicapillary electrophoresis systems using fluorophore labeling with multispectral imaging, to the “next generation” technologies of cyclic array, hybridization based, nanopore and single molecule sequencing. Deciphering the genetic blueprint and follow-up confirmatory sequencing of Homo sapiens and other genomes was only possible by the advent of modern sequencing technologies that was a result of step by step advances with a contribution of academics, medical personnel and instrument companies. While next generation sequencing is moving ahead at break-neck speed, the multicapillary electrophoretic systems played an essential role in the sequencing of the Human Genome, the foundation of the field of genomics. In this prospective, we wish to overview the role of capillary electrophoresis in DNA sequencing based in part of several of our articles in this journal. PMID:19517496

  6. DNA typing by capillary electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, N.

    1997-10-08

    Capillary electrophoresis is becoming more and more important in nucleic acid analysis including DNA sequencing, typing and disease gene measurements. This work summarized the background of DNA typing. The recent development of capillary electrophoresis was also discussed. The second part of the thesis showed the principle of DNA typing based on using the allelic ladder as the absolute standard ladder in capillary electrophoresis system. Future work will be focused on demonstrating DNA typing on multiplex loci and examples of disease diagnosis in the on-line format of PCR-CE. Also capillary array electrophoresis system should allow high throughput, fast speed DNA typing. Only the introduction and conclusions for this report are available here. A reprint was removed for separate processing.

  7. Conducting Polymer Electrodes for Gel Electrophoresis

    OpenAIRE

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

    2014-01-01

    In nearly all cases, electrophoresis in gels is driven via the electrolysis of water at the electrodes, where the process consumes water and produces electrochemical by-products. We have previously demonstrated that p-conjugated polymers such as poly(3,4-ethylenedioxythiophene) (PEDOT) can be placed between traditional metal electrodes and an electrolyte to mitigate electrolysis in liquid (capillary electroosmosis/electrophoresis) systems. In this report, we extend our previous result to gel ...

  8. 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.

  9. Analysis of glycoprotein-derived oligosaccharides in glycoproteins detected on two-dimensional gel by capillary electrophoresis using on-line concentration method.

    Science.gov (United States)

    Kamoda, Satoru; Nakanishi, Yasuharu; Kinoshita, Mitsuhiro; Ishikawa, Rika; Kakehi, Kazuaki

    2006-02-17

    Capillary electrophoresis (CE) is an effective tool to analyze carbohydrate mixture derived from glycoproteins with high resolution. However, CE has a disadvantage that a few nanoliters of a sample solution are injected to a narrow capillary. Therefore, we have to prepare a sample solution of high concentration for CE analysis. In the present study, we applied head column field-amplified sample stacking method to the analysis of N-linked oligosaccharides derived from glycoprotein separated by two-dimensional gel electrophoresis. Model studies demonstrated that we achieved 60-360 times concentration effect on the analysis of carbohydrate chains labeled with 3-aminobenzoic acid (3-AA). The method was applied to the analysis of N-linked oligosaccharides from glycoproteins separated and detected on PAGE gel. Heterogeneity of alpha1-acid glycoprotein (AGP), i.e. glycoforms, was examined by 2D-PAGE and N-linked oligosaccharides were released by in-gel digestion with PNGase F. The released oligosaccharides were derivatized with 3-AA and analyzed by CE. The results showed that glycoforms having lower pI values contained a larger amount of tetra- and tri-antennary oligosaccharides. In contrast, glycoforms having higher pI values contained bi-antennary oligosaccharides abundantly. The result clearly indicated that the spot of a glycoprotein glycoform detected by Coomassie brilliant blue staining on 2D-PAGE gel is sufficient for quantitative profiling of oligosaccharides.

  10. Detection of undeclared animal by-products in commercial canine canned foods: Comparative analyses by ELISA and PCR-RFLP coupled with slab gel electrophoresis or capillary gel electrophoresis.

    Science.gov (United States)

    Hsieh, Ming-Kun; Shih, Pei-Yin; Wei, Chia-Fong; Vickroy, Thomas W; Chou, Chi-Chung

    2016-03-30

    The potential presence of undeclared animal by-products in pet foods is not subject to routine examination. Previously published methods for species-based identification of animal by-products have not been used routinely owing to inconsistent results. The present study evaluated the utility of several approaches for accurate identification of animal by-products in 11 commercial brands of canine canned foods. Canine canned foods from several countries were analysed by ELISA, PCR-RFLP coupled with slab-gel electrophoresis (SGE) and capillary gel electrophoresis (CGE) to test for evidence of by-products derived from cattle, chicken, sheep or pig. While CGE-based analysis detected all (24) animal-derived by-products that were reported for the 11 test samples, SGE and ELISA detected only 22/24 (92%) and 14/24 (58%) of labelled by-products, respectively. In addition, undeclared animal by-products were found using all three analytical approaches with CGE detecting more positives (19) than SGE (17) or ELISA (5). Significant disparities were evident between the labelled contents and the detected content of animal by-products. CGE-based testing for PCR products appears to provide greater sensitivity and accuracy than either SGE or ELISA-based methods. As testing of commercial products becomes more reliable and mainstream, manufacturers will need to develop more thorough and accurate labelling protocols. © 2015 Society of Chemical Industry.

  11. Capillary electrophoresis systems and methods

    Science.gov (United States)

    Dorairaj, Rathissh [Hillsboro, OR; Keynton, Robert S [Louisville, KY; Roussel, Thomas J [Louisville, KY; Crain, Mark M [Georgetown, IN; Jackson, Douglas J [New Albany, IN; Walsh, Kevin M [Louisville, KY; Naber, John F [Goshen, KY; Baldwin, Richard P [Louisville, KY; Franco, Danielle B [Mount Washington, KY

    2011-08-02

    An embodiment of the invention is directed to a capillary electrophoresis apparatus comprising a plurality of separation micro-channels. A sample loading channel communicates with each of the plurality of separation channels. A driver circuit comprising a plurality of electrodes is configured to induce an electric field across each of the plurality of separation channels sufficient to cause analytes in the samples to migrate along each of the channels. The system further comprises a plurality of detectors configured to detect the analytes.

  12. Study of Streptavidin-Modified Quantum Dots by Capillary Electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Stanisavljevic, M.; Janů, L.; Šmerková, K.; Křížková, S.; Pizúrová, Naděžda; Ryvolová, M.; Adam, V.; Hubálek, J.; Kizek, R.

    2013-01-01

    Roč. 76, 7-8 (2013), s. 335-343 ISSN 0009-5893 Institutional support: RVO:68081723 Keywords : Capillary electrophoresis * Gel electrophoresis * Avidin-biotin technology * Oligonucleotide * Nanoparticle * quantum dots Subject RIV: CE - Biochemistry Impact factor: 1.370, year: 2013

  13. Denaturing gradient gel electrophoresis

    International Nuclear Information System (INIS)

    Kocherginskaya, S.A.; Cann, I.K.O.; Mackie, R.I.

    2005-01-01

    It is worthwhile considering that only some 30 species make up the bulk of the bacterial population in human faeces at any one time based on the classical cultivation-based approach. The situation in the rumen is similar. Thus, it is practical to focus on specific groups of interest within the complex community. These may be the predominant or the most active species, specific physiological groups or readily identifiable (genetic) clusters of phylogenetically related organisms. Several 16S rDNA fingerprinting techniques can be invaluable for selecting and monitoring sequences or phylogenetic groups of interest and are described below. Over the past few decades, considerable attention was focussed on the identification of pure cultures of microbes on the basis of genetic polymorphisms of DNA encoding rRNA such as ribotyping, amplified fragment length polymorphism and randomly amplified polymorphic DNA. However, many of these methods require prior cultivation and are less suitable for use in analysis of complex mixed populations although important in describing cultivated microbial diversity in molecular terms. Much less attention was given to molecular characterization of complex communities. In particular, research into diversity and community structure over time has been revolutionized by the advent of molecular fingerprinting techniques for complex communities. Denaturing or temperature gradient gel electrophoresis (DGGE/TGGE) methods have been successfully applied to the analysis of human, pig, cattle, dog and rodent intestinal populations

  14. Detection of clonal immunoglobulin heavy chain gene rearrangements by the polymerase chain reaction and capillary gel electrophoresis.

    Science.gov (United States)

    Fan, Hongxin; Robetorye, Ryan S

    2013-01-01

    Although well-established diagnostic criteria exist for mature B-cell neoplasms, a definitive diagnosis of a B-cell lymphoproliferative disorder cannot always be obtained using more conventional techniques such as flow cytometric immunophenotyping, conventional cytogenetics, fluorescence in situ hybridization, or immunohistochemistry. However, because B-cell malignancies contain identically rearranged immunoglobulin heavy chain genes, the polymerase chain reaction (PCR) can be a fast, convenient, and dependable option to identify clonal B-cell processes. This chapter describes the use of PCR and capillary electrophoresis to identify clonal immunoglobulin heavy chain (IGH) variable and joining region (VH-JH) gene rearrangements (IGH VH-JH PCR) using a commercially available method employing multiple multiplex PCR tubes that was originally developed as the result of a large European BIOMED-2 collaborative study (Invivoscribe Technologies). The core protocol involves the use of three separate master mix tubes that target the conserved framework (FR1, FR2, and FR3) and joining (J) regions of the IGH gene. Analysis of these three framework regions can detect approximately 88% of clonal IGH gene rearrangements.

  15. Microchip capillary gel electrophoresis using programmed field strength gradients for the ultra-fast analysis of genetically modified organisms in soybeans.

    Science.gov (United States)

    Kim, Yun-Jeong; Chae, Joon-Seok; Chang, Jun Keun; Kang, Seong Ho

    2005-08-12

    We have developed a novel method for the ultra-fast analysis of genetically modified organisms (GMOs) in soybeans by microchip capillary gel electrophoresis (MCGE) using programmed field strength gradients (PFSG) in a conventional glass double-T microchip. Under the programmed electric field strength and 0.3% poly(ethylene oxide) sieving matrix, the GMO in soybeans was analyzed within only 11 s of the microchip. The MCGE-PFSG method was a program that changes the electric field strength during GMO analysis, and was also applied to the ultra-fast analysis of PCR products. Compared to MCGE using a conventional and constantly applied electric field, the MCGE-PFSG analysis generated faster results without the loss of resolving power and reproducibility for specific DNA fragments (100- and 250-bp DNA) of GM-soybeans. The MCGE-PFSG technique may prove to be a new tool in the GMO analysis due to its speed, simplicity, and high efficiency.

  16. Capillary array electrophoresis using laser-excited confocal fluorescence detection

    Energy Technology Data Exchange (ETDEWEB)

    Huang, X.C.; Quesada, M.A.; Mathies, R.A. [Univ. of California, Berkeley, CA (United States)

    1992-04-15

    Capillary electrophoresis (CE) has found widespread application in analytical and biomedical research, and the scope and sophistication of CE is still rapidly advancing. Gel-filled capillaries have been employed for the rapid separation and analysis of synthetic polynucleotides, DNA sequencing fragments, and DNA restriction fragments. Open-tube capillary electrophoresis has attained subattomole detection levels in amino acid separations 14 and proven its utility for the separation of proteins, viruses, and bacteria. Separation of the optical isomers of dansyl amino acids has also been successfully demonstrated. Micellar electrokinetic capillary chromatography, isoelectric focusing, and on-column derivatization can all be performed on CE columns, demonstrating the utility of capillary electrophoresis as an analytical and micropreparative tool. 29 refs., 6 figs., 1 tab.

  17. Disc electrophoresis and related techniques of polyacrylamide gel electrophoresis

    National Research Council Canada - National Science Library

    Maurer, H. R

    1971-01-01

    ..., enzymes, antingens and radioactively labelled materials, and detailed treatments of micro disc electrophoresis, preparative polyacrylamide gel electrophoresis and many other techniques for special problems...

  18. Capillary electrophoresis and nanomaterials - Part I: Capillary electrophoresis of nanomaterials.

    Science.gov (United States)

    Adam, Vojtech; Vaculovicova, Marketa

    2017-10-01

    Nanomaterials are in analytical science used for a broad range of purposes, covering the area of sample pretreatment as well as separation, detection, and identification of target molecules. This part of the review covers capillary electrophoresis (CE) of nanomaterials and focuses on the application of CE as a method for characterization used during nanomaterial synthesis and modification as well as the monitoring of their properties and interactions with other molecules. The heterogeneity of the nanomaterial family is extremely large. Depending on different definitions of the term Nanomaterial/Nanoparticle, the group may cover metal and polymeric nanoparticles, carbon nanomaterials, liposomes and even dendrimers. Moreover, these nanomaterials are usually subjected to some kind of surface modification or functionalization, which broadens the diversity even more. Not only for purposes of verification of nanomaterial synthesis and batch-to-batch quality check, but also for determination the polydispersity and for functionality characterization on the nanoparticle surface, has CE offered very beneficial capabilities. Finally, the monitoring of interactions between nanomaterials and other (bio)molecules is easily performed by some kind of capillary electromigration technique. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. Identification and characterization of a thermally cleaved fragment of monoclonal antibody-A detected by sodium dodecyl sulfate-capillary gel electrophoresis.

    Science.gov (United States)

    Kubota, Kei; Kobayashi, Naoki; Yabuta, Masayuki; Ohara, Motomu; Naito, Toyohiro; Kubo, Takuya; Otsuka, Koji

    2017-06-05

    This report describes a novel, comprehensive approach to identifying a fragment peak of monoclonal antibody-A (mAb-A), detected by sodium dodecyl sulfate-capillary gel electrophoresis (SDS-cGE). The fragment migrated close to the internal standard (10kDa marker) of SDS-cGE and increased about 0.5% under a 25°C condition for 6 months. Generally, identification of fragments observed in SDS-cGE is challenging to carry out due to the difficulty of collecting analytical amounts of fractionations from the capillary. In this study, in-gel digestion peptide mapping and reversed phase liquid chromatography-mass spectrometry (RPLC-MS) were employed to elucidate the structure of the fragment. In addition, a Gelfree 8100 fractionation system was newly introduced to collect the fragment and the fraction was applied to the structural analysis of a mAb for the first time. These three analytical methods showed comparable results, proving that the fragment was a fraction of heavy chain HC1-104. The fragment contained complementarity determining regions (CDRs), which are significant to antigen binding, and thus would affect the efficacy of mAb-A. In addition, SDS-cGE without the 10kDa marker was demonstrated to clarify the increased amount of the fragment, and the experiment revealed that the fragment increases 0.2% per year in storage at 5°C. The combination of the three analytical methodologies successfully identified the impurity peak detected by SDS-cGE, providing information critical to assuring the quality and stability of the biotherapeutics. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. Gel electrophoresis of inorganic cations

    International Nuclear Information System (INIS)

    Schoenhofer, F.; Grass, F.

    1978-01-01

    In order to be able to separate the largest possible amounts of substance, polyacryl amide gel (PAA) and silica gel are used as carrier for the electrophoresis. Milligramme quantities can easily be separated on PAA gel plates. Electrophoretic ion focussing considerably improves it. Separations of Sr/Y and lanthanoids were carried out. The behaviour of the readily soluble complexing agent acids on silica gel thin layers was minutely investigated and an interpretation of the focussing effect was derived. The conditions for separating radionuclides were optimized. A further improved separation can be achieved by a time sequence combination of normal electrophoresis and ion focussing. Selective isolation methods are advantageous to determine radionuclide traces in environmental samples. The selective adsorption on preformed deposits was transferred to electrophoresis. After pre-investigations on silica gel layers, strontium and barium could also be retained on PAA gel and radium on strontium sulphate in PAA, whereas the disturbing calcium can easily pass through. Cesium can also be retained by prussian blue in the electrophoresis. (orig.) [de

  1. Separation of intron 22 inversion type 1 and 2 of hemophilia A by modified inverse-shifting polymerase chain reaction and capillary gel electrophoresis.

    Science.gov (United States)

    Pan, Tzu-Yu; Chiou, Shyh-Shin; Wang, Chun-Chi; Wu, Shou-Mei

    2014-12-01

    An inverse-shifting polymerase chain reaction (IS-PCR) combined with short-end capillary gel electrophoresis (CGE) was developed for genotyping of intron 22 inversion Type 1 (Inv22-1) and Type 2 (Inv22-2) of hemophilia A (HA). Severe HA cases are affected by intron 22 inversion around 45-50%. Inv22-1 has higher frequency than Inv22-2. The aim of this study is to distinguish them by genotyping. In order to improve Inv22 genotyping efficiency, five primers were designed and applied to differentiate the wild type, Inv22-1, Inv22-2 and carrier. Three amplicons of 405, 457 and 512 bp were recognized for wild type; 333, 457 and 584 bp for Inv22-1; 385, 405 and 584 bp for Inv22-2. The Inv22-1 carrier has 5 amplicons including 333, 405, 457, 512, 584 bp and Inv22-2 carrier is differentiated by 385, 405, 457, 512 and 584 bp. The amplicons between Inv22-1 and Inv22-2 carriers are only different in 333 bp for Inv22-1 carrier and 385 bp for Inv22-2 carrier. Capillary gel electrophoresis (CGE) was used for separation within 5 min. The separation voltage was set at 8 kV (cathode at detector), and the temperature was kept at 25°C. The sieving matrix was 89 mM Tris, 89 mM boric acid, 2mM EDTA containing 0.4% (w/v) HPMC and 1 μM of YO-PRO(®)-1 Iodide. Total of 50 HA patients (including 35 non-Inv22, 14 Inv22-1, and one Inv22-2 patients) and 7 HA carriers were diagnosed in the application. Seven random samples (5 patients and 2 carriers) were subjected to comparison and gave identical results of DNA sequencing and this modified IS-PCR. Copyright © 2014 Elsevier B.V. All rights reserved.

  2. Characterization of asphaltenes by nonaqueous capillary electrophoresis

    NARCIS (Netherlands)

    Kok, W.T.; Tüdös, A.J.; Grutters, M.; Shepherd, A.G.

    2011-01-01

    Nonaqueous capillary electrophoresis was used for the separation and characterization of asphaltene samples from different sources. For the separation medium (background electrolyte), mixtures of tetrahydrofuran and a high-permittivity organic solvent could be used. The best results were obtained

  3. Analytical biotechnology: Capillary electrophoresis and chromatography

    International Nuclear Information System (INIS)

    Horvath, C.; Nikelly, J.G.

    1990-01-01

    The papers describe the separation, characterization, and equipment required for the electrophoresis or chromatography of cyclic nucleotides, pharmaceuticals, therapeutic proteins, recombinant DNA products, pheromones, peptides, and other biological materials. One paper, On-column radioisotope detection for capillary electrophoresis, has been indexed separately for inclusion on the data base

  4. 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.

  5. DNA gel electrophoresis: the reptation model(s).

    Science.gov (United States)

    Slater, Gary W

    2009-06-01

    DNA gel electrophoresis has been the most important experimental tool to separate DNA fragments for several decades. The introduction of PFGE in the 1980s and capillary gel electrophoresis in the 1990s made it possible to study, map and sequence entire genomes. Explaining how very large DNA molecules move in a gel and why PFGE is needed to separate them has been an active field of research ever since the launch of the journal Electrophoresis. This article presents a personal and historical overview of the development of the theory of gel electrophoresis, focusing on the reptation model, the band broadening mechanisms, and finally the factors that limit the read length and the resolution of electrophoresis-based sequencing systems. I conclude with a short discussion of some of the questions that remain unanswered.

  6. Quantification of Whey Protein Content in Infant Formulas by Sodium Dodecyl Sulfate-Capillary Gel Electrophoresis (SDS-CGE): Single-Laboratory Validation, First Action 2016.15.

    Science.gov (United States)

    Feng, Ping; Fuerer, Christophe; McMahon, Adrienne

    2017-03-01

    Protein separation by sodium dodecyl sulfate-capillary gel electrophoresis, followed by UV absorption at 220 nm, allows for the quantification of major proteins in raw milk. In processed dairy samples such as skim milk powder (SMP) and infant formulas, signals from individual proteins are less resolved, but caseins still migrate as one family between two groups of whey proteins. In the first group, α-lactalbumin and β-lactoglobulin migrate as two distinct peaks. Lactosylated adducts show delayed migration times and interfere with peak separation, but both native and modified forms as well as other low-MW whey proteins still elute before the caseins. The second group contains high-MW whey proteins (including bovine serum albumin, lactoferrin, and immunoglobulins) and elutes after the caseins. Caseins and whey proteins can thus be considered two distinct nonoverlapping families whose ratio can be established based on integrated areas without the need for a calibration curve. Because mass-to-area response factors for whey proteins and caseins are different, an area correction factor was determined from experimental measurement using SMP. Method performance assessed on five infant formulas showed RSDs of 0.2-1.2% (within day) and 0.5-1.1% (multiple days), with average recoveries between 97.4 and 106.4% of added whey protein. Forty-three different infant formulas and milk powders were analyzed. Of the 41 samples with manufacturer claims, the measured whey protein content was in close agreement with declared values, falling within 5% of the declared value in 76% of samples and within 10% in 95% of samples.

  7. Attempt to run urinary protein electrophoresis using capillary technique.

    Science.gov (United States)

    Falcone, Michele

    2014-10-01

    The study of urinary protein has a predominant place in the diagnosis of kidney disease. The most common technique is agarose gel electrophoresis (AGE). For several years, the technique of choice applied to the analysis of serum proteins has been CE, a system that uses capillary fused silica, subjected to high voltage to separate and measure serum proteins. The purpose of this paper was to perform capillary electrophoresis on urinary proteins which, at present, are not interpretable due to the many nonspecific peaks visible when using gel electrophoresis. In order to carry out our research, we used a capillary V8 analyzer together with an agarose gel system from the same company. AGE was taken as the reference method, for which urine was used without any pretreatment. For the V8 system, urine was subjected to purification on granular-activated carbon and then inserted into the V8 analyzer, selecting a program suitable for liquids with low protein content. We examined 19 urine samples collected over 24 hrs from both hospitalized and external patients with different types of proteinuria plus a serum diluted 1/61 considered as a control to recognize the bands. Both methods showed the same protein fractions and classified the proteinuria in a similar way. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Capillaries for use in a multiplexed capillary electrophoresis system

    Science.gov (United States)

    Yeung, E.S.; Chang, H.T.; Fung, E.N.

    1997-12-09

    The invention provides a side-entry optical excitation geometry for use in a multiplexed capillary electrophoresis system. A charge-injection device is optically coupled to capillaries in the array such that the interior of a capillary is imaged onto only one pixel. In Sanger-type 4-label DNA sequencing reactions, nucleotide identification (``base calling``) is improved by using two long-pass filters to split fluorescence emission into two emission channels. A binary poly(ethyleneoxide) matrix is used in the electrophoretic separations. 19 figs.

  9. Capillary Electrophoresis Analysis of Conventional Splicing Assays

    DEFF Research Database (Denmark)

    de Garibay, Gorka Ruiz; Acedo, Alberto; García-Casado, Zaida

    2014-01-01

    of these assays is often challenging. Here, we explore this issue by conducting splicing assays in 31 BRCA2 genetic variants. All variants were assessed by RT-PCR followed by capillary electrophoresis and direct sequencing. If assays did not produce clear-cut outputs (Class-2 or Class-5 according to analytical...

  10. Research on pre-staining gel electrophoresis

    International Nuclear Information System (INIS)

    Zhong Ruibo; Liu Yushuang; Zhang Ping; Liu Jingran; Zhao Guofen; Zhang Feng

    2014-01-01

    Background: Gel electrophoresis is a powerful biochemical separation technique. Most biological molecules are completely transparent in the visible region of light, so it is necessary to use staining to show the results after gel electrophoresis, and the general steps of conventional staining methods are time-consuming. Purpose: We try to develop a novel approach to simplify the gel electrophoresis: Pre-Staining Gel Electrophoresis (PSGE), which can make the gel electrophoresis results monitored in real time. Methods: Pre-stain the protein samples with Coomassie Brilliant Blue (CBB) for 30 min before loading the sample into the gel well. Results and Conclusion: PSGE can be successfully used to analyze the binding efficiency of Bovine Serum Albumin (BSA) and amphiphilic polymer via chemical coupling and physical absorption, and the double PSGE also shows a great potential in bio-analytical chemistry. (authors)

  11. Micro-injector for capillary electrophoresis.

    Science.gov (United States)

    Sáiz, Jorge; Koenka, Israel Joel; García-Ruiz, Carmen; Müller, Beat; Chwalek, Thomas; Hauser, Peter C

    2015-08-01

    A novel micro-injector for capillary electrophoresis for the handling of samples with volumes down to as little as 300 nL was designed and built in our laboratory for analyses in which the available volume is a limitation. The sample is placed into a small cavity located directly in front of the separation capillary, and the injection is then carried out automatically by controlled pressurization of the chamber with compressed air. The system also allows automated flushing of the injection chamber as well as of the capillary. In a trial with a capillary electrophoresis system with contactless conductivity detector, employing a capillary of 25 μm diameter, the results showed good stability of migration times and peak areas. To illustrate the technique, the fast separation of five inorganic cations (Na(+) , K(+) , NH4 (+) , Ca(2+) , and Mg(2+) ) was set up. This could be achieved in less than 3 min, with good limits of detection (10 μM) and linear ranges (between about 10 and 1000 μM). The system was demonstrated for the determination of the inorganic cations in porewater samples of a lake sediment core. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Microfluidic chip-capillary electrophoresis devices

    CERN Document Server

    Fung, Ying Sing; Du, Fuying; Guo, Wenpeng; Ma, Tongmei; Nie, Zhou; Sun, Hui; Wu, Ruige; Zhao, Wenfeng

    2015-01-01

    Capillary electrophoresis (CE) and microfluidic chip (MC) devices are relatively mature technologies, but this book demonstrates how they can be integrated into a single, revolutionary device that can provide on-site analysis of samples when laboratory services are unavailable. By introducing the combination of CE and MC technology, Microfluidic Chip-Capillary Electrophoresis Devices broadens the scope of chemical analysis, particularly in the biomedical, food, and environmental sciences. The book gives an overview of the development of MC and CE technology as well as technology that now allows for the fabrication of MC-CE devices. It describes the operating principles that make integration possible and illustrates some achievements already made by the application of MC-CE devices in hospitals, clinics, food safety, and environmental research. The authors envision further applications for private and public use once the proof-of-concept stage has been passed and obstacles to increased commercialization are ad...

  13. Recent applications of nanomaterials in capillary electrophoresis.

    Science.gov (United States)

    González-Curbelo, Miguel Ángel; Varela-Martínez, Diana Angélica; Socas-Rodríguez, Bárbara; Hernández-Borges, Javier

    2017-10-01

    Nanomaterials have found an important place in Analytical Chemistry and, in particular, in Separation Science. Among them, metal-organic frameworks, magnetic and non-magnetic nanoparticles, carbon nanotubes and graphene, as well as their combinations, are the most important nanomaterials that have been used up to now. Concerning capillary electromigration techniques, these nanomaterials have also been used as both pseudostationary phases in electrokinetic chromatography (EKC) and as stationary phases in microchip capillary electrophoresis (CE) and capillary electrochromatography (CEC), as a result of their interesting and particular properties. This review article pretends to provide a general and critical revision of the most recent applications of nanomaterials in this field (period 2010-2017). © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. Usage of capillary electrophoresis for common hemoglobinopathies screening

    Directory of Open Access Journals (Sweden)

    Alireza Ebrahimi

    2016-06-01

    Full Text Available Hemoglobinopathies are most common inherited disorders in the world; approximately 7 percent of the worldwide population and 5-6 percent of population of Iran are carriers. The hemoglobin disorders inherit as autosomal recessive and are very common in the Mediterranean area and much of the Asia and Africa. The control of this inherited disorders need to genetic counseling and accurate screening by more advanced and more accurate methods. This study explains features of current Iran hemoglobin disorders, nominates the accessible methods for screening them and introduces the capillary zone electrophoresis as a rapid and more accurate method. The required data were extracted of various articles and then for good explanation, current Iran hemoglobinopathies properties were showed in the tables and electropherograms of important hemoglobin disorders in Iran population were provided for help to interpretation results of blood tests by capillary zone electrophoresis method. Hemoglobin disorders are including thalassemias and hemoglobin variants; Disruption in the production and malfunction of globin chains cause types of hemoglobin disorders. We cannot introduce one of clinical laboratory tests as critical and basic method for screening and distinguishing types of inherited hemoglobin disorders as alone. For distinguishing the types of them must be prepared enough information and data of the hemoglobin disorders and for more accurate analysis must be used simultaneously different methods as gel electrophoresis, high performance liquid chromatography, isoelectric focusing, capillary zone electrophoresis or molecular tests. The capillary electrophoresis is an accurate and rapid method for screening types of the hemoglobin disorders. Other side this method cannot analyze all of them, so must be used biochemical, biophysical and molecular methods for confirmation the results. This review showed we can use the capillary electrophoresis and HPLC as two

  15. Usage of Capillary Electrophoresis for screening common Hemoglobinopathies

    Directory of Open Access Journals (Sweden)

    2016-06-01

    Full Text Available Hemoglobinopathies are most common inherited disorders in the world approximately 7 percent of the worldwide population and 5-6 percent of population of Iran are carriers. For control of this inherited hemoglobin disorders need to accurate screening by more advanced and more accurate methods. This study explains features of current Iran hemoglobin disorders, nominates the accessible methods for screening them and introduces the capillary zone electrophoresis as a rapid & more accurate method. The required data were extracted of various articles and then for good explanation, current Iran hemoglobinopathies properties were showed in the tables and electropherograms of important hemoglobin disorders in Iran population were provided for help to interpretation results of blood tests by capillary zone electrophoresis method. Hemoglobin disorders are including thalassemias & hemoglobin variants Disruption in the production and malfunction of globin chains cause types of hemoglobin disorders. We cannot introduce one of clinical laboratory tests as critical and basic method for screening and distinguishing types of inherited hemoglobin disorders as alone. For distinguishing the types of them must be prepared enough information and data of the hemoglobin disorders and for more accurate analysis must be used simultaneously different methods as Gel electrophoresis, High performance liquid chromatography, Isoelectric focusing, Capillary zone electrophoresis or molecular tests. The capillary electrophoresis is an accurate and rapid method for screening types of the hemoglobin disorders. Other side this method cannot analyze all of them, so must be used biochemical, biophysical and molecular methods for confirmation the results. This review showed we can use the capillary electrophoresis and HPLC as two complementary methods for hemoglobinopathies screening. We can analyze by the methods more hemoglobin disorders and decrease more laboratory errors. Moreover

  16. Van de Graaff generator for capillary electrophoresis.

    Science.gov (United States)

    Lee, Seung Jae; Castro, Eric R; Guijt, Rosanne M; Tarn, Mark D; Manz, Andreas

    2017-09-29

    A new approach for high voltage capillary electrophoresis (CE) is proposed, which replaces the standard high voltage power supply with a Van de Graaff generator, a low current power source. Because the Van de Graaff generator is a current-limited source (10μA), potentials exceeding 100kV can be generated for CE when the electrical resistance of the capillary is maximized. This was achieved by decreasing the capillary diameter and reducing the buffer ionic strength. Using 2mM borate buffer and a 5μm i.d. capillary, fluorescently labeled amino acids were separated with efficiencies up to 3.5 million plates; a 5.7 fold improvement in separation efficiency compared to a normal power supply (NPS) typically used in CE. This separation efficiency was realized using a simple set-up without significant Joule heating, making the Van de Graaff generator a promising alternative for applying the high potentials required for enhancing resolution in the separation and analysis of highly complex samples, for example mixtures of glycans. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. Sheathless interface for coupling capillary electrophoresis with mass spectrometry

    Science.gov (United States)

    Wang, Chenchen; Tang, Keqi; Smith, Richard D.

    2014-06-17

    A sheathless interface for coupling capillary electrophoresis (CE) with mass spectrometry is disclosed. The sheathless interface includes a separation capillary for performing CE separation and an emitter capillary for electrospray ionization. A portion of the emitter capillary is porous or, alternatively, is coated to form an electrically conductive surface. A section of the emitter capillary is disposed within the separation capillary, forming a joint. A metal tube, containing a conductive liquid, encloses the joint.

  18. Detection of clonal T-cell receptor beta and gamma chain gene rearrangement by polymerase chain reaction and capillary gel electrophoresis.

    Science.gov (United States)

    Fan, Hongxin; Robetorye, Ryan S

    2013-01-01

    Although established diagnostic criteria exist for mature T-cell neoplasms, a definitive diagnosis of a T-cell lymphoproliferative disorder cannot always be obtained using more conventional techniques such as flow cytometric immunophenotyping, conventional cytogenetics, fluorescence in situ hybridization, or immunohistochemistry. However, because T-cell malignancies contain identically rearranged T-cell receptor gamma (TCRG) and/or beta (TCRB) genes, the polymerase chain reaction (PCR) can be a fast, convenient, and dependable option to identify clonal T-cell processes. This chapter describes the use of PCR and capillary electrophoresis to identify clonal TCRB and TCRG gene rearrangements (TCRB and TCRG PCR) using a commercially available method employing multiple multiplex PCR tubes that was originally developed as the result of a large European BIOMED-2 collaborative study (Invivoscribe Technologies). The core protocol for the TCRB assay involves the use of three separate multiplex master mix tubes. Tubes A and B target the framework regions within the variable and joining regions of the TCRB gene, and Tube C targets the diversity and joining regions of the TCRB gene. The core protocol for the TCRG assay utilizes two multiplex master mix tubes (Tubes A and B) that target the variable and joining regions of the TCRG gene. Use of the five BIOMED-2 TCRB and TCRG PCR multiplex tubes in parallel can detect approximately 94% of clonal TCR gene rearrangements.

  19. Large abnormal peak on capillary zone electrophoresis due to contrast agent.

    Science.gov (United States)

    Wheeler, Rachel D; Zhang, Liqun; Sheldon, Joanna

    2017-01-01

    Background Some iodinated radio-contrast media absorb ultraviolet light and can therefore be detected by capillary zone electrophoresis. If seen, these peaks are typically small with 'quantifications' of below 5 g/L. Here, we describe the detection of a large peak on capillary zone electrophoresis that was due to the radio-contrast agent, Omnipaque™. Methods Serum from a patient was analysed by capillary zone electrophoresis, and the IgG, IgA, IgM and total protein concentrations were measured. The serum sample was further analysed by gel electrophoresis and immunofixation. Results Capillary zone electrophoresis results for the serum sample showed a large peak with a concentration high enough to warrant urgent investigation. However, careful interpretation alongside the serum immunoglobulin concentrations and total protein concentration showed that the abnormal peak was a pseudoparaprotein rather than a monoclonal immunoglobulin. This was confirmed by analysis with gel electrophoresis and also serum immunofixation. The patient had had a CT angiogram with the radio-contrast agent Omnipaque™; addition of Omnipaque™ to a normal serum sample gave a peak with comparable mobility to the pseudoparaprotein in the patient's serum. Conclusions Pseudoparaproteins can appear as a large band on capillary zone electrophoresis. This case highlights the importance of a laboratory process that detects significant electrophoretic abnormalities promptly and interprets them in the context of the immunoglobulin concentrations. This should avoid incorrect reporting of pseudoparaproteins which could result in the patient having unnecessary investigations.

  20. Supramolecular gel electrophoresis of large DNA fragments.

    Science.gov (United States)

    Tazawa, Shohei; Kobayashi, Kazuhiro; Oyoshi, Takanori; Yamanaka, Masamichi

    2017-10-01

    Pulsed-field gel electrophoresis is a frequent technique used to separate exceptionally large DNA fragments. In a typical continuous field electrophoresis, it is challenging to separate DNA fragments larger than 20 kbp because they migrate at a comparable rate. To overcome this challenge, it is necessary to develop a novel matrix for the electrophoresis. Here, we describe the electrophoresis of large DNA fragments up to 166 kbp using a supramolecular gel matrix and a typical continuous field electrophoresis system. C 3 -symmetric tris-urea self-assembled into a supramolecular hydrogel in tris-boric acid-EDTA buffer, a typical buffer for DNA electrophoresis, and the supramolecular hydrogel was used as a matrix for electrophoresis to separate large DNA fragments. Three types of DNA marker, the λ-Hind III digest (2 to 23 kbp), Lambda DNA-Mono Cut Mix (10 to 49 kbp), and Marker 7 GT (10 to 165 kbp), were analyzed in this study. Large DNA fragments of greater than 100 kbp showed distinct mobility using a typical continuous field electrophoresis system. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Potential of capillary electrophoresis for the profiling of propolis

    NARCIS (Netherlands)

    Hilhorst, M.J; Somsen, G.W; de Jong, G.J.

    1998-01-01

    The usefulness of capillary electrophoresis (CE) with diode array detection for the profiling of Propolis, a hive product, is investigated. Water extracts of Propolis were analyzed with both capillary zone electrophoresis (CZE) at pH 7.0 and 9.3, and micellar electrokinetic chromatography (MEKC)

  2. Quantitative analysis by microchip capillary electrophoresis – current limitations and problem-solving strategies

    NARCIS (Netherlands)

    Revermann, T.; Götz, S.; Künnemeyer, Jens; Karst, U.

    2008-01-01

    Obstacles and possible solutions for the application of microchip capillary electrophoresis in quantitative analysis are described and critically discussed. Differences between the phenomena occurring during conventional capillary electrophoresis and microchip-based capillary electrophoresis are

  3. Capillaries modified by noncovalent anionic polymer adsorption for capillary zone electrophoresis, micellar electrokinetic capillary chromatography and capillary electrophoresis mass spectrometry

    DEFF Research Database (Denmark)

    Bendahl, L; Hansen, S H; Gammelgaard, Bente

    2001-01-01

    A simple coating procedure for generation of a high and pH-independent electroosmotic flow in capillary zone electrophoresis (CZE) and micellar electrokinetic capillary chromatography (MEKC) is described. The bilayer coating was formed by noncovalent adsorption of the ionic polymers Polybrene...... capillaries was (4.9+/-0.1) x 10(-4) cm2V(-1)s(-1) in a pH-range of 2-10 (ionic strength = 30 mM). When alkaline compounds were used as test substances intracapillary and intercapillary migration time variations (n = 6) were less than 1% relative standard deviation (RSD) and 2% RSD, respectively in the entire...... pH range. The coating was fairly stable in the presence of sodium dodecyl sulfate, and this made it possible to perform fast MEKC separations at low pH. When neutral compounds were used as test substances, the intracapillary migration time variations (n = 6) were less than 2% RSD in a pH range of 2...

  4. DNA DAMAGE QUANTITATION BY ALKALINE GEL ELECTROPHORESIS.

    Energy Technology Data Exchange (ETDEWEB)

    SUTHERLAND,B.M.; BENNETT,P.V.; SUTHERLAND, J.C.

    2004-03-24

    Physical and chemical agents in the environment, those used in clinical applications, or encountered during recreational exposures to sunlight, induce damages in DNA. Understanding the biological impact of these agents requires quantitation of the levels of such damages in laboratory test systems as well as in field or clinical samples. Alkaline gel electrophoresis provides a sensitive (down to {approx} a few lesions/5Mb), rapid method of direct quantitation of a wide variety of DNA damages in nanogram quantities of non-radioactive DNAs from laboratory, field, or clinical specimens, including higher plants and animals. This method stems from velocity sedimentation studies of DNA populations, and from the simple methods of agarose gel electrophoresis. Our laboratories have developed quantitative agarose gel methods, analytical descriptions of DNA migration during electrophoresis on agarose gels (1-6), and electronic imaging for accurate determinations of DNA mass (7-9). Although all these components improve sensitivity and throughput of large numbers of samples (7,8,10), a simple version using only standard molecular biology equipment allows routine analysis of DNA damages at moderate frequencies. We present here a description of the methods, as well as a brief description of the underlying principles, required for a simplified approach to quantitation of DNA damages by alkaline gel electrophoresis.

  5. Acetic acid denaturing pulsed field capillary electrophoresis for RNA separation.

    Science.gov (United States)

    Li, Zhenqing; Dou, Xiaoming; Ni, Yi; Sumitomo, Keiko; Yamaguchi, Yoshinori

    2010-10-01

    Based on our previous work of in-capillary denaturing polymer electrophoresis, we present a study of RNA molecular separation up to 6.0 kilo nucleotide by pulsed field CE. This is the first systematic investigation of electrophoresis of a larger molecular mass RNA in linear hydroxyethylcellulose (HEC) under pulsed field conditions. The parameters that may influence the separation performance, e.g. gel polymer concentration, modulation depth and pulse frequency, are analyzed in terms of resolution and mobility. For denaturing and separating RNA in the capillary simultaneously, 2 M acetic acid was added into the HEC polymer to serve as separation buffer. Result shows that (i) in pulsed field conditions, RNA separation can be achieved in a wide range of concentration of HEC polymer, and RNA fragments between 0.3 and 0.6 kilo nucleotide are sensitive to the polymer concentration; (ii) under certain pulsed field conditions, RNA fragments move linearly as the modulation depth increases; (iii) 12.5 Hz is the resonance frequency for RNA reorientation time and applied frequency.

  6. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS ...

    African Journals Online (AJOL)

    Four strains of eri, Samia cynthia ricini Lepidoptera: Saturniidae that can be identified morphologically and maintained at North East Institute of Science and Technology, Jorhat were characterized based on their protein profile by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and DNA by random ...

  7. Matching Two-dimensional Gel Electrophoresis' Spots

    DEFF Research Database (Denmark)

    Dos Anjos, António; AL-Tam, Faroq; Shahbazkia, Hamid Reza

    2012-01-01

    This paper describes an approach for matching Two-Dimensional Electrophoresis (2-DE) gels' spots, involving the use of image registration. The number of false positive matches produced by the proposed approach is small, when compared to academic and commercial state-of-the-art approaches. This ar...

  8. Pulsed field gel electrophoresis a practical guide

    CERN Document Server

    Birren, Bruce

    1993-01-01

    Pulsed Field Gel Electrophoresis: A Practical Guide is the first laboratory manual to describe the theory and practice of this technique. Based on the authors' experience developing pulsed field gel instruments and teaching procedures, this book provides everything a researcher or student needs to know in order to understand and carry out pulsed field gel experiments. Clear, well-tested protocols assume only that users have a basic familiarity with molecular biology. Thorough coverage of useful data, theory, and applications ensures that this book is also a lasting resource for more adv

  9. Capillary electrophoresis: principles and applications in illicit drug analysis.

    Science.gov (United States)

    Tagliaro, F; Turrina, S; Smith, F P

    1996-02-09

    Capillary electrophoresis, which appeared in the early 1980s, is now rapidly expanding into many scientific disciplines, including analytical chemistry, biotechnology and biomedical and pharmaceutical sciences. In capillary electrophoresis,electrokinetic separations are carried out in tiny capillaries at high voltages (10-30 kV), thus obtaining high efficiencies (N > 10(5)) and excellent mass sensitivities (down to 10(-18)-10(-20) moles). The main features of capillary electrophoresis are: versatility of application (from inorganic ions to large DNA fragments), use of different separation modes with different selectivity, extremely low demands on sample volume, negligible running costs, possibility of interfacing with different detection systems, ruggedness and simplicity of instrumentation. Capillary electrophoresis applications in forensic sciences have appeared only recently, but are now rapidly growing, particularly in forensic toxicology. The present paper briefly describes the basic principles of capillary electrophoresis, from both the instrumental and analytical points of view. Furthermore, the main applications in the analysis of illicit/controlled drugs in both illicit preparations and biological samples are presented and discussed (43 references). It is concluded that the particular separation mechanism and the high complementarity of this technique to chromatography makes capillary electrophoresis a new powerful tool of investigation in the hands of forensic toxicologists.

  10. Two-dimensional capillary electrophoresis: capillary isoelectric focusing and capillary zone electrophoresis with laser-induced fluorescence detection

    Science.gov (United States)

    Dickerson, Jane A.; Ramsay, Lauren M.; Dada, Oluwatosin O.; Cermak, Nathan

    2011-01-01

    Capillary isoelectric focusing and capillary zone electrophoresis are coupled with laser-induced fluorescence detection to create an ultrasensitive two-dimensional separation method for proteins. In this method, two capillaries are joined through a buffer filled interface. Separate power supplies control the potential at the injection end of the first capillary and at the interface; the detector is held at ground potential. Proteins are labeled with the fluorogenic reagent Chromeo P503, which preserves the isoelectric point of the labeled protein. The labeled proteins were mixed with ampholytes and injected into the first dimension capillary. A focusing step was performed with the injection end of the capillary at high pH and the interface at low pH. To mobilize components, the interface was filled with a high pH buffer, which was compatible with the second dimension separation. A fraction was transferred to the second dimension capillary for separation. The process of fraction transfer and second dimension separation was repeated two dozen times. The separation produced a spot capacity of 125. PMID:20603830

  11. Fabricating PFPE Membranes for Capillary Electrophoresis

    Science.gov (United States)

    Lee, Michael C.; Willis, Peter A.; Greer, Frank; Rolland, Jason

    2009-01-01

    A process has been developed for fabricating perfluoropolyether (PFPE) membranes that contain microscopic holes of precise sizes at precise locations. The membranes are to be incorporated into laboratory-on-a-chip microfluidic devices to be used in performing capillary electrophoresis. The present process is a modified version of part of the process, described in the immediately preceding article, that includes a step in which a liquid PFPE layer is cured into solid (membrane) form by use of ultraviolet light. In the present process, one exploits the fact that by masking some locations to prevent exposure to ultraviolet light, one can prevent curing of the PFPE in those locations. The uncured PFPE can be washed away from those locations in the subsequent release and cleaning steps. Thus, holes are formed in the membrane in those locations. The most straightforward way to implement the modification is to use, during the ultraviolet-curing step, an ultraviolet photomask similar to the photomasks used in fabricating microelectronic devices. In lieu of such a photomask, one could use a mask made of any patternable ultraviolet-absorbing material (for example, an ink or a photoresist).

  12. TECHNIQUES WITH POTENTIAL FOR HANDLING ENVIRONMENTAL SAMPLES IN CAPILLARY ELECTROPHORESIS

    Science.gov (United States)

    An assessment of the methods for handling environmental samples prior to capillary electrophoresis (CE) is presented for both aqueous and solid matrices. Sample handling in environmental analyses is the subject of ongoing research at the Environmental Protection Agency's National...

  13. Capillary electrophoresis in the N-glycosylation analysis of biopharmaceuticals

    Czech Academy of Sciences Publication Activity Database

    Guttman, András

    2013-01-01

    Roč. 48, JUL-AUG (2013), s. 132-143 ISSN 0165-9936 Institutional support: RVO:68081715 Keywords : automated workflow * biopharmaceuticals * capillary electrophoresis Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 6.612, year: 2013

  14. Success and failure with phthalate buffers in capillary zone electrophoresis

    NARCIS (Netherlands)

    Bocek, P.; Gebauer, P.; Beckers, J.L.

    2001-01-01

    Phthalate buffers are currently used in capillary electrophoresis as robust electrolyte systems for indirect detection. This contribution demonstrates that these buffers show regularly not only successful regions of mobilities of analytes (sample window) but also regions of failure where the

  15. Methods and instrumentation for quantitative microchip capillary electrophoresis

    NARCIS (Netherlands)

    Revermann, T.

    2007-01-01

    The development of novel instrumentation and analytical methodology for quantitative microchip capillary electrophoresis (MCE) is described in this thesis. Demanding only small quantities of reagents and samples, microfluidic instrumentation is highly advantageous. Fast separations at high voltages

  16. Field-portable Capillary Electrophoresis Instrument with Conductivity Detection

    International Nuclear Information System (INIS)

    Zhang, H F; Liu, X W; Wang, W; Wang, X L; Tian, L

    2006-01-01

    In this paper a novel capillary electrophoresis chip (CEC) is presented with integrated platinum electrodes and simplified conductivity detector. CEC is fabricated by the method of mechanical modification with probe on organic glass. Capillary electrophoresis chip can rapidly completed ion separation by simulation of concentration distribution and zone-broadening. Detection circuit is simple which can detect pA order current. This system has those advantages such as small volume, low power consumption and linearity, and well suit for field analysis

  17. Performing Isoelectric Focusing and Simultaneous Fractionation of Proteins on A Rotary Valve Followed by Sodium Dodecyl – Polyacrylamide Gel Electrophoresis

    Science.gov (United States)

    Wang, Wei; Lu, Joann J.; Gu, Congying; Zhou, Lei; Liu, Shaorong

    2013-01-01

    In this technical note, we design and fabricate a novel rotary valve and demonstrate its feasibility for performing isoelectric focusing and simultaneous fractionation of proteins, followed by sodium dodecyl – polyacrylamide gel electrophoresis. The valve has two positions. In one position, the valve routes a series of capillary loops together into a single capillary tube where capillary isoelectric focusing (CIEF) is performed. By switching the valve to another position, the CIEF-resolved proteins in all capillary loops are isolated simultaneously, and samples in the loops are removed and collected in vials. After the collected samples are briefly processed, they are separated via sodium dodecyl – polyacrylamide gel electrophoresis (SDS-PAGE, the 2nd-D separation) on either a capillary gel electrophoresis instrument or a slab-gel system. The detailed valve configuration is illustrated, and the experimental conditions and operation protocols are discussed. PMID:23819755

  18. Development of high-throughput and high sensitivity capillary gel electrophoresis platform method for Western, Eastern, and Venezuelan equine encephalitis (WEVEE) virus like particles (VLPs) purity determination and characterization.

    Science.gov (United States)

    Gollapudi, Deepika; Wycuff, Diane L; Schwartz, Richard M; Cooper, Jonathan W; Cheng, K C

    2017-10-01

    In this paper, we describe development of a high-throughput, highly sensitive method based on Lab Chip CGE-SDS platform for purity determination and characterization of virus-like particle (VLP) vaccines. A capillary gel electrophoresis approach requiring about 41 s per sample for analysis and demonstrating sensitivity to protein initial concentrations as low as 20 μg/mL, this method has been used previously to evaluate monoclonal antibodies, but this application for lot release assay of VLPs using this platform is unique. The method was qualified and shown to be accurate for the quantitation of VLP purity. Assay repeatability was confirmed to be less than 2% relative standard deviation of the mean (% RSD) with interday precision less than 2% RSD. The assay can evaluate purified VLPs in a concentration range of 20-249 μg/mL for VEE and 20-250 μg/mL for EEE and WEE VLPs. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. Simultaneous detection of genetically modified organisms by multiplex ligation-dependent genome amplification and capillary gel electrophoresis with laser-induced fluorescence.

    Science.gov (United States)

    García-Cañas, Virginia; Mondello, Monica; Cifuentes, Alejandro

    2010-07-01

    In this work, an innovative method useful to simultaneously analyze multiple genetically modified organisms is described. The developed method consists in the combination of multiplex ligation-dependent genome dependent amplification (MLGA) with CGE and LIF detection using bare-fused silica capillaries. The MLGA process is based on oligonucleotide constructs, formed by a universal sequence (vector) and long specific oligonucleotides (selectors) that facilitate the circularization of specific DNA target regions. Subsequently, the circularized target sequences are simultaneously amplified with the same couple of primers and analyzed by CGE-LIF using a bare-fused silica capillary and a run electrolyte containing 2-hydroxyethyl cellulose acting as both sieving matrix and dynamic capillary coating. CGE-LIF is shown to be very useful and informative for optimizing MLGA parameters such as annealing temperature, number of ligation cycles, and selector probes concentration. We demonstrate the specificity of the method in detecting the presence of transgenic DNA in certified reference and raw commercial samples. The method developed is sensitive and allows the simultaneous detection in a single run of percentages of transgenic maize as low as 1% of GA21, 1% of MON863, and 1% of MON810 in maize samples with signal-to-noise ratios for the corresponding DNA peaks of 15, 12, and 26, respectively. These results demonstrate, to our knowledge for the first time, the great possibilities of MLGA techniques for genetically modified organisms analysis.

  20. Capillary Electrophoresis Artifact Due to Eosin

    Science.gov (United States)

    Murphy, Kathleen M.; Berg, Karin D.; Geiger, Tanya; Hafez, Michael; Flickinger, Katie A.; Cooper, Lisa; Pearson, Patrick; Eshleman, James R.

    2005-01-01

    Capillary electrophoresis (CE) is a commonly used tool in the analysis of fluorescently labeled PCR amplification products. We have identified a CE artifact caused by the tissue stain eosin that can complicate the interpretation of CE data. The artifact was detected during routine analysis of a DNA sample isolated from a formalin-fixed, paraffin-embedded tissue sample considered histologically suspicious for a B-cell neoplasm. A standard clinical PCR and CE assay for immunoglobulin heavy chain (IGH) gene rearrangement revealed a weak polyclonal population of rearranged IGH genes and a 71 base peak suspicious for IGH clonality. The spectral properties of the 71 base peak were unusual in that although the dominant fluorescence of the peak was blue, it also fluoresced in green and yellow (blue>green>yellow), raising the suspicion that the peak might represent an artifact. CE analysis of the genomic DNA sample without PCR amplification demonstrated the presence of the 71 base peak, suggesting that the artifact was caused by a contaminant within the DNA sample itself. We demonstrate that eosin, which was used to stain the formalin-fixed tissue during processing, yields a discrete 71 base peak of similar morphology to the contaminant peak on CE analysis. The data suggest that eosin in the fixed tissue was not completely eliminated during nucleic acid extraction, resulting in the artifact peak. We discuss the implications of this potentially common contaminant on the interpretation of CE data and demonstrate that artifacts caused by eosin can be avoided by using more stringent DNA purification steps. Histological dyes may fluoresce, and artifacts from them should be considered when primary peaks contain additional underlying peaks of other colors. PMID:15681487

  1. Integration of amperometric sensors for microchip capillary electrophoresis application

    International Nuclear Information System (INIS)

    Dicorato, F; Moore, E; Glennon, J

    2011-01-01

    Capillary electrophoresis is a technique for the separation and analysis of chemical compounds. Techniques adopted from the microchip technology knowledge have led to recent developments of electrophoresis system with integration on microchip. Microchip Capillary Electrophoresis (μCE) systems offer a series of advantages as easy integration for Lab-on-a-chip applications, high performance, portability, speed, minimal solvent and sample requirements. A new technological challenge aims at the development of an economic modular microchip capillary electrophoresis systems using separable and independent units concerning the sensor. In this project we worked on the development of an interchangeable amperometric sensor in order to provide a solution to such electrode passivation and facilitating the use of tailored sensors for specific analyte detection besides. Fluidic chips have been machined from cyclic olefin polymer pallets (Zeonor) using a micro-injection molding machine.

  2. Integration of amperometric sensors for microchip capillary electrophoresis application

    Energy Technology Data Exchange (ETDEWEB)

    Dicorato, F; Moore, E [Life Sciences Interface Group, Tyndall National Institute, Lee Maltings, Dyke Parade, Cork (Ireland); Glennon, J, E-mail: eric.moore@tyndall.ie [Chemistry Department, University College Cork, College Road, Cork (Ireland)

    2011-08-17

    Capillary electrophoresis is a technique for the separation and analysis of chemical compounds. Techniques adopted from the microchip technology knowledge have led to recent developments of electrophoresis system with integration on microchip. Microchip Capillary Electrophoresis ({mu}CE) systems offer a series of advantages as easy integration for Lab-on-a-chip applications, high performance, portability, speed, minimal solvent and sample requirements. A new technological challenge aims at the development of an economic modular microchip capillary electrophoresis systems using separable and independent units concerning the sensor. In this project we worked on the development of an interchangeable amperometric sensor in order to provide a solution to such electrode passivation and facilitating the use of tailored sensors for specific analyte detection besides. Fluidic chips have been machined from cyclic olefin polymer pallets (Zeonor) using a micro-injection molding machine.

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

    International Nuclear Information System (INIS)

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

    2003-01-01

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

  4. An integrated multiple capillary array electrophoresis system for high-throughput DNA sequencing

    Energy Technology Data Exchange (ETDEWEB)

    Lu, X.

    1998-03-27

    A capillary array electrophoresis system was chosen to perform DNA sequencing because of several advantages such as rapid heat dissipation, multiplexing capabilities, gel matrix filling simplicity, and the mature nature of the associated manufacturing technologies. There are two major concerns for the multiple capillary systems. One concern is inter-capillary cross-talk, and the other concern is excitation and detection efficiency. Cross-talk is eliminated through proper optical coupling, good focusing and immersing capillary array into index matching fluid. A side-entry excitation scheme with orthogonal detection was established for large capillary array. Two 100 capillary array formats were used for DNA sequencing. One format is cylindrical capillary with 150 {micro}m o.d., 75 {micro}m i.d and the other format is square capillary with 300 {micro}m out edge and 75 {micro}m inner edge. This project is focused on the development of excitation and detection of DNA as well as performing DNA sequencing. The DNA injection schemes are discussed for the cases of single and bundled capillaries. An individual sampling device was designed. The base-calling was performed for a capillary from the capillary array with the accuracy of 98%.

  5. Using Gel Electrophoresis To Illustrate Protein Diversity and Isoelectric Point.

    Science.gov (United States)

    Browning, Mark; Vanable, Joseph

    2002-01-01

    Demonstrates the differences in protein structures by focusing on isoelectric point with an experiment that is observable under certain pH levels in gel electrophoresis. Explains the electrophoresis procedure and reports results of the experiments. (YDS)

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

    Science.gov (United States)

    Anderson, James; Wright, Drew; Meksem, Khalid

    2013-01-01

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

  7. Recent advances of capillary electrophoresis in pharmaceutical analysis.

    Science.gov (United States)

    Suntornsuk, Leena

    2010-09-01

    This review covers recent advances of capillary electrophoresis (CE) in pharmaceutical analysis. The principle, instrumentation, and conventional modes of CE are briefly discussed. Advances in the different CE techniques (non-aqueous CE, microemulsion electrokinetic chromatography, capillary isotachophoresis, capillary electrochromatography, and immunoaffinity CE), detection techniques (mass spectrometry, light-emitting diode, fluorescence, chemiluminescence, and contactless conductivity), on-line sample pretreatment (flow injection) and chiral separation are described. Applications of CE to assay of active pharmaceutical ingredients (APIs), drug impurity testing, chiral drug separation, and determination of APIs in biological fluids published from 2008 to 2009 are tabulated.

  8. Capillary electrophoresis-based assessment of nanobody affinity and purity

    NARCIS (Netherlands)

    Haselberg, Rob; Oliveira, Sabrina; van der Meel, Roy; Somsen, Govert W; de Jong, Gerhardus J

    2014-01-01

    Drug purity and affinity are essential attributes during development and production of therapeutic proteins. In this work, capillary electrophoresis (CE) was used to determine both the affinity and composition of the biotechnologically produced "nanobody" EGa1, the binding fragment of a

  9. Capillary zone electrophoresis-mass spectromet of intact proteins

    NARCIS (Netherlands)

    Domínguez-Vega, Elena; Haselberg, Rob; Somsen, Govert W.

    2016-01-01

    Capillary electrophoresis (CE) coupled with mass spectrometry (MS) has proven to be a powerful analytical tool for the characterization of intact proteins. It combines the high separation efficiency, short analysis time, and versatility of CE with the mass selectivity and sensitivity offered by MS

  10. Application of a diode-array detector in capillary electrophoresis

    NARCIS (Netherlands)

    Beck, W.; Hoek, van R.; Engelhardt, H.

    1993-01-01

    In the last decade diode-array detection has proved to be extremely useful in high performance liquid chromatography in recording UV-visible spectra directly and on-line in the column effluent. In capillary electrophoresis (CE) only fast-scanning detectors with long scan times (up to 2 s) are

  11. Thermostatted dual-channel portable capillary electrophoresis instrument

    Czech Academy of Sciences Publication Activity Database

    Koenka, I.J.; Küng, N.; Kubáň, Pavel; Chwalek, T.; Furrer, G.; Wehrli, B.; Müller, B.; Hauser, P.C.

    2016-01-01

    Roč. 37, 17-18 (2016), s. 2368-2375 ISSN 0173-0835 Institutional support: RVO:68081715 Keywords : portable devices * on-site measurements * capillary electrophoresis Subject RIV: CB - Analytical Chemistry , Separation Impact factor: 2.744, year: 2016

  12. Characterization of metal/humic acid systems by Capillary Electrophoresis

    NARCIS (Netherlands)

    Staden JJ van; Hoop MAGT van den; Cleven R; LAC

    2000-01-01

    Metal-humic acid systems have been characterised applying Capillary Electrophoresis (CE). Appropriate experimental conditions with respect to carrier electrolyte, pH range, salt concentration, humic acid concentration and the applied potential, have been optimised. The influence of multivalent metal

  13. Capillary electrophoresis in the analysis of biologically important thiols

    Czech Academy of Sciences Publication Activity Database

    Lačná, J.; Kubáň, Petr; Foret, František

    2017-01-01

    Roč. 38, č. 1 (2017), s. 203-222 ISSN 0173-0835 Institutional support: RVO:68081715 Keywords : biological thiols * capillary electrophoresis * clinical applications Subject RIV: CB - Analytical Chemistry, Separation OBOR OECD: Analytical chemistry Impact factor: 2.744, year: 2016

  14. Thermostatted dual-channel portable capillary electrophoresis instrument

    Czech Academy of Sciences Publication Activity Database

    Koenka, I.J.; Küng, N.; Kubáň, Pavel; Chwalek, T.; Furrer, G.; Wehrli, B.; Müller, B.; Hauser, P.C.

    2016-01-01

    Roč. 37, 17-18 (2016), s. 2368-2375 ISSN 0173-0835 Institutional support: RVO:68081715 Keywords : portable devices * on-site measurements * capillary electrophoresis Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 2.744, year: 2016

  15. Determination of propionate in bread using capillary zone electrophoresis

    NARCIS (Netherlands)

    Ackermans, M.T.; Ackermans-Loonen, J.C.J.M; Beckers, J.L.

    1992-01-01

    A method for the determination of propionate in bread is described. The propionate was extracted from the bread with a repeated extraction procedure and measured using capillary zone electrophoresis in the indirect UV mode applying a background electrolyte of 0.005 M Tris adjusted at pH 4.6 by

  16. Comparison of non-electrophoresis grade with electrophoresis grade BIS in NIPAM polymer gel preparation

    Science.gov (United States)

    Khodadadi, Roghayeh; Khajeali, Azim; Farajollahi, Ali Reza; Hajalioghli, Parisa; Raeisi, Noorallah

    2015-01-01

    Introduction:The main objective of this study was to investigate the possibility of replacing electrophoresis cross-linker with non-electrophoresis N, N′-methylenebisacrylamide (BIS) in N-isopropyl acrylamide (NIPAM) polymer gel and its possible effect on dose response. Methods: NIPAM polymer gel was prepared from non-electrophoresis grade BIS and the relaxation rate (R2) was measured by MR imaging after exposing the gel to gamma radiation from Co-60 source. To compare the response of this gel with the one that contains electrophoresis grade BIS, two sets of NIPAM gel were prepared using electrophoresis and non-electrophoresis BIS and irradiated to different gamma doses. Results: It was found that the dose–response of NIPAM gel made from the non-electrophoresis grade BIS is coincident with that of electrophoresis grade BIS. Conclusion:Taken all, it can be concluded that the non-electrophoresis grade BIS not only is a suitable alternative for the electrophoresis grade BIS but also reduces the cost of gel due to its lower price. PMID:26457250

  17. The gel electrophoresis markup language (GelML) from the Proteomics Standards Initiative.

    Science.gov (United States)

    Gibson, Frank; Hoogland, Christine; Martinez-Bartolomé, Salvador; Medina-Aunon, J Alberto; Albar, Juan Pablo; Babnigg, Gyorgy; Wipat, Anil; Hermjakob, Henning; Almeida, Jonas S; Stanislaus, Romesh; Paton, Norman W; Jones, Andrew R

    2010-09-01

    The Human Proteome Organisation's Proteomics Standards Initiative has developed the GelML (gel electrophoresis markup language) data exchange format for representing gel electrophoresis experiments performed in proteomics investigations. The format closely follows the reporting guidelines for gel electrophoresis, which are part of the Minimum Information About a Proteomics Experiment (MIAPE) set of modules. GelML supports the capture of metadata (such as experimental protocols) and data (such as gel images) resulting from gel electrophoresis so that laboratories can be compliant with the MIAPE Gel Electrophoresis guidelines, while allowing such data sets to be exchanged or downloaded from public repositories. The format is sufficiently flexible to capture data from a broad range of experimental processes, and complements other PSI formats for MS data and the results of protein and peptide identifications to capture entire gel-based proteome workflows. GelML has resulted from the open standardisation process of PSI consisting of both public consultation and anonymous review of the specifications.

  18. Inexpensive and Safe DNA Gel Electrophoresis Using Household Materials

    Science.gov (United States)

    Ens, S.; Olson, A. B.; Dudley, C.; Ross, N. D., III; Siddiqi, A. A.; Umoh, K. M.; Schneegurt, M. A.

    2012-01-01

    Gel electrophoresis is the single most important molecular biology technique and it is central to life sciences research, but it is often too expensive for the secondary science classroom or homeschoolers. A simple safe low-cost procedure is described here that uses household materials to construct and run DNA gel electrophoresis. Plastic…

  19. An improved interface for capillary zone electrophoresis-mass spectrometry

    International Nuclear Information System (INIS)

    Smith, R.D.; Loo, J.A.; Barinaga, C.J.; Udseth, H.R.

    1988-06-01

    We have recently developed an improved electrospray ionization interface for capillary electrophoresis mass-spectrometry (CZE-MS). Our initial interface employed a vacuum deposited metal film at the exit of the capillary to make an electrical contact with he eluting buffer and establish the electrospray field gradient. This interface did, however, impose significant limitations on the range of capillary electrophoretic (CE) separations that could be performed. To circumvent these limitations, an interface that does not require a metalized tip was designed nd developed. In the new approach, the electrical contact at the column exit is made through a flowing liquid sheath. The principal advantage of this interface is that it allows operation with a much broader range of electrophoresis conditions. The sheath flow can be readily varied in both composition and volume. An electrospray ionization spectrum is given for a previously intractable buffer solution. 5 refs., 2 figs

  20. Integrated refractive index optical ring resonator detector for capillary electrophoresis.

    Science.gov (United States)

    Zhu, Hongying; White, Ian M; Suter, Jonathan D; Zourob, Mohammed; Fan, Xudong

    2007-02-01

    We developed a novel miniaturized and multiplexed, on-capillary, refractive index (RI) detector using liquid core optical ring resonators (LCORRs) for future development of capillary electrophoresis (CE) devices. The LCORR employs a glass capillary with a diameter of approximately 100 mum and a wall thickness of a few micrometers. The circular cross section of the capillary forms a ring resonator along which the light circulates in the form of the whispering gallery modes (WGMs). The WGM has an evanescent field extending into the capillary core and responds to the RI change due to the analyte conducted in the capillary, thus permitting label-free measurement. The resonating nature of the WGM enables repetitive light-analyte interaction, significantly enhancing the LCORR sensitivity. This LCORR architecture achieves dual use of the capillary as a sensor head and a CE fluidic channel, allowing for integrated, multiplexed, and noninvasive on-capillary detection at any location along the capillary. In this work, we used electro-osmotic flow and glycerol as a model system to demonstrate the fluid transport capability of the LCORRs. In addition, we performed flow speed measurement on the LCORR to demonstrate its flow analysis capability. Finally, using the LCORR's label-free sensing mechanism, we accurately deduced the analyte concentration in real time at a given point on the capillary. A sensitivity of 20 nm/RIU (refractive index units) was observed, leading to an RI detection limit of 10-6 RIU. The LCORR marries photonic technology with microfluidics and enables rapid on-capillary sample analysis and flow profile monitoring. The investigation in this regard will open a door to novel high-throughput CE devices and lab-on-a-chip sensors in the future.

  1. An analytical model for enantioseparation process in capillary electrophoresis

    Science.gov (United States)

    Ranzuglia, G. A.; Manzi, S. J.; Gomez, M. R.; Belardinelli, R. E.; Pereyra, V. D.

    2017-12-01

    An analytical model to explain the mobilities of enantiomer binary mixture in capillary electrophoresis experiment is proposed. The model consists in a set of kinetic equations describing the evolution of the populations of molecules involved in the enantioseparation process in capillary electrophoresis (CE) is proposed. These equations take into account the asymmetric driven migration of enantiomer molecules, chiral selector and the temporary diastomeric complexes, which are the products of the reversible reaction between the enantiomers and the chiral selector. The solution of these equations gives the spatial and temporal distribution of each species in the capillary, reproducing a typical signal of the electropherogram. The mobility, μ, of each specie is obtained by the position of the maximum (main peak) of their respective distributions. Thereby, the apparent electrophoretic mobility difference, Δμ, as a function of chiral selector concentration, [ C ] , can be measured. The behaviour of Δμ versus [ C ] is compared with the phenomenological model introduced by Wren and Rowe in J. Chromatography 1992, 603, 235. To test the analytical model, a capillary electrophoresis experiment for the enantiomeric separation of the (±)-chlorpheniramine β-cyclodextrin (β-CD) system is used. These data, as well as, other obtained from literature are in closed agreement with those obtained by the model. All these results are also corroborate by kinetic Monte Carlo simulation.

  2. Analysis of glycated hemoglobin A1c by capillary electrophoresis and capillary isoelectric focusing

    Czech Academy of Sciences Publication Activity Database

    Koval, Dušan; Kašička, Václav; Cottet, H.

    2011-01-01

    Roč. 413, č. 1 (2011), s. 8-15 ISSN 0003-2697 R&D Projects: GA ČR GP203/09/P485; GA ČR(CZ) GA203/08/1428 Institutional research plan: CEZ:AV0Z40550506 Keywords : capillary zone electrophoresis * capillary isoelectric focusing * glycated hemoglobin HbA1c Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 2.996, year: 2011

  3. Capillary electrophoresis microchip coupled with on-line chemiluminescence detection

    International Nuclear Information System (INIS)

    Su Rongguo; Lin Jinming; Qu Feng; Chen Zhifeng; Gao Yunhua; Yamada, Masaaki

    2004-01-01

    In the present work, chemiluminescence detection was integrated with capillary electrophoresis microchip. The microchip was designed on the principle of flow-injection chemiluminescence system and capillary electrophoresis. It has three main channels, five reservoirs and a detection cell. As model samples, dopamine and catechol were separated and detected using a permanganate chemiluminescent system on the prepared microchip. The samples were electrokinetically injected into the double-T cross section, separated in the separation channel, and then oxidized by chemiluminescent reagent delivered by a home-made micropump to produce light in the detection cell. The electroosmotic flow could be smoothly coupled with the micropump flow. The detection limits for dopamine and catechol were 20.0 and 10.0 μM, respectively. Successful separation and detection of dopamine and catechol demonstrated the distinct advantages of integration of chemiluminescent detection on a microchip for rapid and sensitive analysis

  4. DNA migration mechanism analyses for applications in capillary and microchip electrophoresis

    Science.gov (United States)

    Forster, Ryan E.; Hert, Daniel G.; Chiesl, Thomas N.; Fredlake, Christopher P.; Barron, Annelise E.

    2009-01-01

    In 2009, electrophoretically driven DNA separations in slab gels and capillaries have the sepia tones of an old-fashioned technology in the eyes of many, even while they remain ubiquitously used, fill a unique niche, and arguably have yet to reach their full potential. For comic relief, what is old becomes new again: agarose slab gel separations are used to prepare DNA samples for “next-gen” sequencing platforms (e.g., the Illumina and 454 machines)—dsDNA molecules within a certain size range are “cut out” of a gel and recovered for subsequent “massively parallel” pyrosequencing. In this review, we give a Barron lab perspective on how our comprehension of DNA migration mechanisms in electrophoresis has evolved, since the first reports of DNA separations by CE (∼1989) until now, 20 years later. Fused silica capillaries, and borosilicate glass and plastic microchips, quietly offer increasing capacities for fast (and even “ultra-fast”), efficient DNA separations. While the channel-by-channel scaling of both old and new electrophoresis platforms provides key flexibility, it requires each unique DNA sample to be prepared in its own micro- or nanovolume. This Achille's heel of electrophoresis technologies left an opening through which pooled-sample, next-gen DNA sequencing technologies rushed. We shall see, over time, whether sharpening understanding of transitions in DNA migration modes in crosslinked gels, nanogel solutions, and uncrosslinked polymer solutions will allow electrophoretic DNA analysis technologies to flower again. Microchannel electrophoresis, after a quiet period of metamorphosis, may emerge sleeker and more powerful, to claim its own important niche applications. PMID:19582705

  5. Quantification of nucleotides by ICPMS: coupling of ICPMS with capillary electrophoresis or capillary HPLC

    International Nuclear Information System (INIS)

    Inagaki, K.; Fujii, S.; Takatsu, A.; Yarita, T.; Zhu, Y.; Chiba, K.

    2009-01-01

    Full text: Quantification of nucleotides in small volumes of biological samples has eagerly been demanded. A method using ICPMS coupled with capillary electrophoresis or capillary liquid chromatography is reported. A new interface system, which consists of a double tube nebulizer inserted with a fused silica capillary tube and a cylinder mini-chamber with a sheath gas inlet, was designed. Moreover, the surface conditions of the sampling and skimmer cones, and the introduction of H 2 gas into the plasma were found to significantly improve the signal/background ratio for phosphorus determination at m/z 31. (author)

  6. Comprehensive protein profiling by multiplexed capillary zone electrophoresis using cross-linked polyacrylamide coated capillaries.

    Science.gov (United States)

    Liu, Shaorong; Gao, Lin; Pu, Qiaosheng; Lu, Joann J; Wang, Xingjia

    2006-02-01

    We have recently developed a new process to create cross-linked polyacrylamide (CPA) coatings on capillary walls to suppress protein-wall interactions. Here, we demonstrate CPA-coated capillaries for high-efficiency (>2 x 10(6) plates per meter) protein separations by capillary zone electrophoresis (CZE). Because CPA virtually eliminates electroosmotic flow, positive and negative proteins cannot be analyzed in a single run. A "one-sample-two-separation" approach is developed to achieve a comprehensive protein analysis. High throughput is achieved through a multiplexed CZE system.

  7. Capillary electrophoresis in a fused-silica capillary with surface roughness gradient

    Czech Academy of Sciences Publication Activity Database

    Horká, Marie; Šlais, Karel; Karásek, Pavel; Růžička, F.; Šalplachta, Jiří; Šesták, Jozef; Kahle, Vladislav; Roth, Michal

    2016-01-01

    Roč. 39, č. 19 (2016), s. 3827-3834 ISSN 1615-9306 R&D Projects: GA ČR(CZ) GA16-03749S; GA MZd(CZ) NV16-29916A Institutional support: RVO:68081715 Keywords : capillary electrophoresis * supercritical water * surface roughness gradient Subject RIV: CB - Analytical Chemistry , Separation Impact factor: 2.557, year: 2016

  8. Capillary electrophoresis in a fused-silica capillary with surface roughness gradient

    Czech Academy of Sciences Publication Activity Database

    Horká, Marie; Šlais, Karel; Karásek, Pavel; Růžička, F.; Šalplachta, Jiří; Šesták, Jozef; Kahle, Vladislav; Roth, Michal

    2016-01-01

    Roč. 39, č. 19 (2016), s. 3827-3834 ISSN 1615-9306 R&D Projects: GA ČR(CZ) GA16-03749S; GA MZd(CZ) NV16-29916A Institutional support: RVO:68081715 Keywords : capillary electrophoresis * supercritical water * surface roughness gradient Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 2.557, year: 2016

  9. Gel Electrophoresis on a Budget to Dye for

    Science.gov (United States)

    Yu, Julie H.

    2010-01-01

    Gel electrophoresis is one of the most important tools used in molecular biology and has facilitated the entire field of genetic engineering by enabling the separation of nucleic acids and proteins. However, commercial electrophoresis kits can cost up to $800 for each setup, which is cost prohibitive for most classroom budgets. This article…

  10. Characterization and Study of Transgenic Cultivars by Capillary and Microchip Electrophoresis

    Directory of Open Access Journals (Sweden)

    Elena Domínguez Vega

    2014-12-01

    Full Text Available Advances in biotechnology have increased the demand for suitable analytical techniques for the analysis of genetically modified organisms. Study of the substantial equivalence, discrimination between transgenic and non-transgenic cultivars, study of the unintended effects caused by a genetic modification or their response to diverse situations or stress conditions (e.g., environmental, climatic, infections are some of the concerns that need to be addressed. Capillary electrophoresis (CE is emerging as an alternative to conventional techniques for the study and characterization of genetically modified organisms. This article reviews the most recent applications of CE for the analysis and characterization of transgenic cultivars in the last five years. Different strategies have been described depending on the level analyzed (DNA, proteins or metabolites. Capillary gel electrophoresis (CGE has shown to be particularly useful for the analysis of DNA fragments amplified by PCR. Metabolites and proteins have been mainly separated using capillary zone electrophoresis (CZE using UV and MS detection. Electrophoretic chips have also proven their ability in the analysis of transgenic cultivars and a section describing the new applications is also included.

  11. Analysis of lipoproteins by capillary zone electrophoresis in microfluidic devices: Assay development and surface roughness measurements

    NARCIS (Netherlands)

    Weiller, Bruce H.; Ceriotti, Laura; Shibata, Takayuki; Rein, Dietrich; Roberts, Matthew A.; Lichtenberg, Jan; German, J. Bruce; De Rooij, Nico F.; Verpoorte, Elisabeth

    2002-01-01

    The development of a new assay for lipoproteins by capillary electrophoresis in fused-silica capillaries and in glass microdevices is described in this paper. The separation of low-density (LDL) and high-density (HDL) lipoproteins by capillary zone electrophoresis is demonstrated in fused-silica

  12. An air-pressure-free elastomeric valve for integrated nucleic acid analysis by capillary electrophoresis

    International Nuclear Information System (INIS)

    Jung, Wooseok; Barrett, Matthew; Brooks, Carla; Zenhausern, Frederic; Rivera, Andrew; Birdsell, Dawn N; Wagner, David M

    2015-01-01

    We present a new elastomeric valve for integrated nucleic acid analysis by capillary electrophoresis. The valve functions include metering to capture a designated volume of biological sample into a polymerase chain reaction (PCR) chamber, sealing to preserve the sample during PCR cycling, and transfer of the PCR-products and on-chip formamide post-processing for the analysis of DNA fragments by capillary gel electrophoresis. This new valve differs from prior art polydimethylsiloxane (PDMS) valves in that the valve is not actuated externally by air-pressure or vacuum so that it simplifies a DNA analysis system by eliminating the need for an air-pressure or vacuum source, and off-cartridge solenoid valves, control circuit boards and software. Instead, the new valve is actuated by a thermal cycling peltier assembly integrated within the hardware instrument that tightly comes in contact with a microfluidic cartridge for thermal activation during PCR, so that it spontaneously closes the valve without an additional actuator system. The valve has bumps in the designated locations so that it has a self-alignment that does not require precise alignment of a valve actuator. Moreover, the thickness of the new valve is around 600 μm with an additional bump height of 400 μm so that it is easy to handle and very feasible to fabricate by injection molding compared to other PDMS valves whose thicknesses are around 30–100 μm. The new valve provided over 95% of metering performance in filling the fixed volume of the PCR chamber, preserved over 97% of the sample volume during PCR, and showed very comparable capillary electrophoresis peak heights to the benchtop assay tube controls with very consistent transfer volume of the PCR-product and on-chip formamide. The new valve can perform a core function for integrated nucleic acid analysis by capillary electrophoresis. (paper)

  13. Development of novel separation techniques for biological samples in capillary electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Chang, Huan -Tsung [Iowa State Univ., Ames, IA (United States)

    1994-07-27

    This dissertation includes three different topics: general introduction of capillary electrophoresis (CE); gradient in CE and CE in biological separations; and capillary gel electrophoresis (CGE) for DNA separation. Factors such as temperature, viscosity, pH, and the surface of capillary walls affecting the separation performance are demonstrated. A pH gradient between 3.0 and 5.2 is useful to improve the resolution among eight different organic acids. A flow gradient due to the change in the concentration of surfactant, which is able to coat to the capillary wall to change the flow rate and its direction, is also shown as a good way to improve the resolution for organic compounds. A temperature gradient caused by joule heat is shown by voltage programming to enhance the resolution and shorten the separation time for several phenolic compounds. The author also shows that self-regulating dynamic control of electroosmotic flow in CE by simply running separation in different concentrations of surfactant has less matrix effect on the separation performance. One of the most important demonstrations in this dissertation is that the author proposes on-column reaction which gives several advantages including the use of a small amount of sample, low risk of contamination, and time saving and kinetic features. The author uses this idea with laser induced fluorescence (LIF) as a detection mode to detect an on-column digestion of sub-ng of protein. This technique also is applied to single cell analysis in the group.

  14. Ceramic protective coatings applied by sol-gel or electrophoresis

    International Nuclear Information System (INIS)

    Stoch, A.

    1993-01-01

    Sol-gel and electrophoresis are the complementary techniques which may be used for obtaining the ceramic coatings. The composition of such a coatings depends on the composition of electrophoresis bath or sol solution. Thermal treatment is used for densifying the coating and promoting the adherence of coating to the substrate. In presented work silica, silica-alumina or alumina coatings are applied by sol-gel dip coating procedure on steel, aluminium or ceramic substrates. Electrophoresis is employed for obtaining zirconia, alumina or hydroxyapatite coatings on stainless steel. (author). 7 refs

  15. Capillary Electrophoresis Sensitivity Enhancement Based on Adaptive Moving Average Method.

    Science.gov (United States)

    Drevinskas, Tomas; Telksnys, Laimutis; Maruška, Audrius; Gorbatsova, Jelena; Kaljurand, Mihkel

    2018-06-05

    In the present work, we demonstrate a novel approach to improve the sensitivity of the "out of lab" portable capillary electrophoretic measurements. Nowadays, many signal enhancement methods are (i) underused (nonoptimal), (ii) overused (distorts the data), or (iii) inapplicable in field-portable instrumentation because of a lack of computational power. The described innovative migration velocity-adaptive moving average method uses an optimal averaging window size and can be easily implemented with a microcontroller. The contactless conductivity detection was used as a model for the development of a signal processing method and the demonstration of its impact on the sensitivity. The frequency characteristics of the recorded electropherograms and peaks were clarified. Higher electrophoretic mobility analytes exhibit higher-frequency peaks, whereas lower electrophoretic mobility analytes exhibit lower-frequency peaks. On the basis of the obtained data, a migration velocity-adaptive moving average algorithm was created, adapted, and programmed into capillary electrophoresis data-processing software. Employing the developed algorithm, each data point is processed depending on a certain migration time of the analyte. Because of the implemented migration velocity-adaptive moving average method, the signal-to-noise ratio improved up to 11 times for sampling frequency of 4.6 Hz and up to 22 times for sampling frequency of 25 Hz. This paper could potentially be used as a methodological guideline for the development of new smoothing algorithms that require adaptive conditions in capillary electrophoresis and other separation methods.

  16. Pulsed-field gel electrophoresis of bacterial chromosomes.

    Science.gov (United States)

    Mawer, Julia S P; Leach, David R F

    2013-01-01

    The separation of fragments of DNA by agarose gel electrophoresis is integral to laboratory life. Nevertheless, standard agarose gel electrophoresis cannot resolve fragments bigger than 50 kb. Pulsed-field gel electrophoresis is a technique that has been developed to overcome the limitations of standard agarose gel electrophoresis. Entire linear eukaryotic chromosomes, or large fragments of a chromosome that have been generated by the action of rare-cutting restriction endonucleases, can be separated using this technique. As a result, pulsed-field gel electrophoresis has many applications, from karyotype analysis of microbial genomes, to the analysis of chromosomal strand breaks and their repair intermediates, to the study of DNA replication and the identification of origins of replication. This chapter presents a detailed protocol for the preparation of Escherichia coli chromosomal DNA that has been embedded in agarose plugs, digested with the rare-cutting endonuclease NotI, and separated by contour-clamped homogeneous field electrophoresis. The principles in this protocol can be applied to the separation of all fragments of DNA whose size range is between 40 kb and 1 Mb.

  17. Gel Electrophoresis--The Easy Way for Students

    Science.gov (United States)

    VanRooy, Wilhelmina; Sultana, Khalida

    2010-01-01

    This article describes a simple, inexpensive, easy to conduct gel-electrophoresis activity using food dyes. It is an alternative to the more expensive counterparts which require agarose gel, DNA samples, purchased chamber and Tris-borate-EDTA buffer. We suggest some learning activities for senior biology students along with comments on several…

  18. Evaluation of wheat by polyacrylamide gel electrophoresis | Shuaib ...

    African Journals Online (AJOL)

    ... polyacrylamide gel electrophoresis (SDS-PAGE). Electrophorogram for each variety were scored and presence or absence of each band noted and was entered in a binary data matrix. Based on the data of SDS-PAGE gels cluster analysis was performed to check the variations among varieties. The overall result shows ...

  19. A simple gel electrophoresis method for separating polyhedral gold nanoparticles

    Science.gov (United States)

    Kim, Suhee; Lee, Hye Jin

    2015-07-01

    In this paper, a simple approach to separate differently shaped and sized polyhedral gold nanoparticles (NPs) within colloidal solutions via gel electrophoresis is described. Gel running parameters for separating efficiently gold NPs including gel composition, added surfactant types and applied voltage were investigated. The plasmonic properties and physical structure of the separated NPs extracted from the gel matrix were then investigated using transmission electron microscopy (TEM) and UV-vis spectrophotometry respectively. Data analysis revealed that gel electrophoresis conditions of a 1.5 % agarose gel with 0.1 % sodium dodecyl sulfate (SDS) surfactant under an applied voltage of 100 V resulted in the selective isolation of ~ 50 nm polyhedral shaped gold nanoparticles. Further efforts are underway to apply the method to purify biomolecule-conjugated polyhedral Au NPs that can be readily used for NP-enhanced biosensing platforms.

  20. Determination of antibacterial flomoxef in serum by capillary electrophoresis.

    Science.gov (United States)

    Kitahashi, Toshihiro; Furuta, Itaru

    2003-04-01

    A determination method of flomoxef (FMOX) concentration in serum by capillary electrophoresis is developed. Serum samples are extracted with acetonitrile. After pretreatment, they are separated in a fused-silica capillary tube with a 25 mM borate buffer (pH 10.0) as a running buffer that contains 50mM sodium dodecyl sulfate. The FMOX and acetaminophen (internal standard) are detected by UV absorbance at 200 nm. Linearity (0-200 mg/L) is good, and the minimum limit of detection is 1.0 mg/L (S/N = 3). The relative standard deviations of intra- and interassay variability are 1.60-4.78% and 2.10-3.31%, respectively, and the recovery rate is 84-98%. This method can be used for determination of FMOX concentration in serum.

  1. Stacking and discontinuous buffers in capillary zone electrophoresis.

    Science.gov (United States)

    Shihabi, Z K

    2000-08-01

    Discontinuous buffers for capillary zone electrophoresis (CZE) can be used under less rigid conditions compared to those for isotachophoresis for stacking. They can be prepared simply by modifying the sample itself, either by addition of small inorganic ions, low conductivity diluents, or both, and also by adjusting its pH, meanwhile injecting a large volume on the capillary. Zwitterionic and organic-based buffers such as triethanolamine and tris(hydroxymethyl)aminomethane (Tris) are well suited for stacking due to their low conductivity, provided the buffer is discontinuous as demonstrated here. A simple mechanism based on discontinuous buffers is described to explain many of the observed stacking types in CZE, pointing out the many similarities to transient isotachophoresis.

  2. Separation and determination of some carboxylic acids by capillary electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Sladkov, V.; Fourest, B

    2006-07-01

    Separation and determination of some organic acids, mono-carboxylic (formic and acetic), dicarboxylic (oxalic and tartaric), tricarboxylic (citric) acids and aromatic acids (phtalic, benzoic, mellitic and trimellitic), by capillary electrophoresis are reviewed. The method development parameters, such as separation and injection mode, are discussed. Special attention is paid to the comparison of different detection types (spectroscopic and electrochemical). The optimisation of the carrier electrolyte composition (choice of carrier electrolyte, effect of pH, ionic strength, electro-osmotic flow modifier) is treated. Different additives (alkali-earth and transition metal ions, cyclodextrins and alcohol), which are often used for improving organic acid separation, are also considered. (authors)

  3. Multivalent weak electrolytes - risky background electrolytes for capillary zone electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Beckers, J. L.; Boček, Petr

    2002-01-01

    Roč. 23, č. 12 (2002), s. 1942-1946 ISSN 0173-0835 R&D Projects: GA ČR GA203/99/0044; GA ČR GA203/02/0023; GA ČR GA203/01/0401; GA AV ČR IAA4031703; GA AV ČR IAA4031103 Institutional research plan: CEZ:AV0Z4031919 Keywords : background electrolytes * capillary zone electrophoresis * multivalent electrolytes Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 4.325, year: 2002

  4. Separation and determination of some carboxylic acids by capillary electrophoresis

    International Nuclear Information System (INIS)

    Sladkov, V.; Fourest, B.

    2006-01-01

    Separation and determination of some organic acids, mono-carboxylic (formic and acetic), dicarboxylic (oxalic and tartaric), tricarboxylic (citric) acids and aromatic acids (phtalic, benzoic, mellitic and trimellitic), by capillary electrophoresis are reviewed. The method development parameters, such as separation and injection mode, are discussed. Special attention is paid to the comparison of different detection types (spectroscopic and electrochemical). The optimisation of the carrier electrolyte composition (choice of carrier electrolyte, effect of pH, ionic strength, electro-osmotic flow modifier) is treated. Different additives (alkali-earth and transition metal ions, cyclodextrins and alcohol), which are often used for improving organic acid separation, are also considered. (authors)

  5. Separation and quantification of cellulases and hemicellulases by capillary electrophoresis

    DEFF Research Database (Denmark)

    Jørgensen, Henning; Kutter, Jörg Peter; Olsson, Lisbeth

    2003-01-01

    Cellulases and hemicellulases are two classes of enzymes produced by filamentous fungi and secreted into the cultivation medium. Both classes of enzymes consist of a subset of classes of which the fungi produce several enzymes with varying molecular mass and pI but similar enzymatic activities....... Current methods are limited in their ability to quantify all of these enzymes when all are present simultaneously in a mixture. Five different cellulases (two cellobiohydrolases and three endoglucanases) and one hemicellulase (endoxylanase) were separated using capillary electrophoresis (CE) in a fused...

  6. Capillary electrophoresis-MALDI interface based on inkjet technology

    Science.gov (United States)

    Vannatta, Michael W.; Whitmore, Colin D.; Dovichi, Norman J.

    2010-01-01

    An ink jet printer valve and nozzle were used to deliver matrix and sample from an electrophoresis capillary onto a MALDI plate. The system was evaluated by separation of a set of standard peptides. That separation generated up to 40,000 theoretical plates in less than three minutes. Detection limits were 500 amol using an ABI TOF-TOF instrument and 2 fmol for an ABI Q-TOF instrument. Over 70% coverage was obtained for the tryptic digest of α-lactalbumin in less than 2.5 minutes. PMID:19960472

  7. Ergot alkaloids as chiral selectors in capillary electrophoresis and other electromigration methods

    Czech Academy of Sciences Publication Activity Database

    Sinibaldi, M.; Messina, A.; Stodůlková, Eva; Flieger, Miroslav

    2010-01-01

    Roč. 1, č. 3 (2010), s. 233-243 ISSN 0976-5514 Institutional research plan: CEZ:AV0Z50200510 Keywords : capillary electrophoresis * capillary electrochromatography * chiral analysis Subject RIV: CB - Analytical Chemistry, Separation

  8. Agarose gel electrophoresis for the separation of DNA fragments.

    Science.gov (United States)

    Lee, Pei Yun; Costumbrado, John; Hsu, Chih-Yuan; Kim, Yong Hoon

    2012-04-20

    Agarose gel electrophoresis is the most effective way of separating DNA fragments of varying sizes ranging from 100 bp to 25 kb(1). Agarose is isolated from the seaweed genera Gelidium and Gracilaria, and consists of repeated agarobiose (L- and D-galactose) subunits(2). During gelation, agarose polymers associate non-covalently and form a network of bundles whose pore sizes determine a gel's molecular sieving properties. The use of agarose gel electrophoresis revolutionized the separation of DNA. Prior to the adoption of agarose gels, DNA was primarily separated using sucrose density gradient centrifugation, which only provided an approximation of size. To separate DNA using agarose gel electrophoresis, the DNA is loaded into pre-cast wells in the gel and a current applied. The phosphate backbone of the DNA (and RNA) molecule is negatively charged, therefore when placed in an electric field, DNA fragments will migrate to the positively charged anode. Because DNA has a uniform mass/charge ratio, DNA molecules are separated by size within an agarose gel in a pattern such that the distance traveled is inversely proportional to the log of its molecular weight(3). The leading model for DNA movement through an agarose gel is "biased reptation", whereby the leading edge moves forward and pulls the rest of the molecule along(4). The rate of migration of a DNA molecule through a gel is determined by the following: 1) size of DNA molecule; 2) agarose concentration; 3) DNA conformation(5); 4) voltage applied, 5) presence of ethidium bromide, 6) type of agarose and 7) electrophoresis buffer. After separation, the DNA molecules can be visualized under uv light after staining with an appropriate dye. By following this protocol, students should be able to: Understand the mechanism by which DNA fragments are separated within a gel matrix Understand how conformation of the DNA molecule will determine its mobility through a gel matrix Identify an agarose solution of appropriate

  9. Rapid monitoring of autolysis process of proteases by capillary electrophoresis.

    Science.gov (United States)

    Chen, Xiu-Lan; Shun, Cai-Yun; Zhang, Yu-Zhong; Gao, Pei-Ji

    2003-10-01

    A protease, MCP-01, produced by a deep-sea psychrotrophic strain of Pseudoaltermonas sp. SM9913 was purified and its autolysis reaction at 20 degrees C-50 degrees C was monitored by capillary electrophoresis. Capillary electrophoresis provides a rapid assay because the degree and state of autolysis of protease MCP-01 could be observed within 6 min. The autolysis rate increased as the temperature rose in the tested range. After 30 min incubation at 30 degrees C, 77% of MCP-01 autolyzed into peptides. However, its activity for the hydrolysis of casein was reduced by only 4%. The rate of loss of activity of MCP-01 was thus slower than that of autolysis of MCP-01 at 30 degrees C. Similar results were obtained when MCP-01 was incubated at 20 degrees C, 40 degrees C and 50 degrees C. Large peptides produced by autolysis of MCP-01 therefore still have catalytic activity. When these large peptides autolyzed further into smaller peptides, the enzyme conformation that retained its catalytic activity was destroyed and activity was lost.

  10. Success and failure with phthalate buffers in capillary zone electrophoresis.

    Science.gov (United States)

    Bocek, P; Gebauer, P; Beckers, J L

    2001-04-01

    Phthalate buffers are currently used in capillary electrophoresis as robust electrolyte systems for indirect detection. This contribution demonstrates that these buffers show regularly not only successful regions of mobilities of analytes (sample window) but also regions of failure where the migration of analytes is strongly deteriorated due to the presence of a system zone. System zones in phthalate buffers may be easily detected by UV detection and manifest themselves as peaks or dips. Peak shape diagrams are advantageously used for the prediction of the migration behavior of system zones in phthalate background electrolyte (BGE) systems at various pH. It is shown that the mobility of the system zone varies strongly with pH, is practically zero at pH values below 4 and above 7, and shows a maximum at pH 5. Thus, the system peak may coincide either with the peaks of various analytes or with the electroosmotic flow (EOF) peak. Experiments are given showing the effects of such coincidences as, e.g., zigzag detection patterns, double EOF peaks, and/or unusually broad peaks/dips. The message of this contribution is to show how to understand the electrophoretic properties of phthalate BGEs that, regardless of possible failure regions, may be successfully used in the analytical practice of capillary zone electrophoresis (CZE).

  11. Analytical characterization of wine and its precursors by capillary electrophoresis.

    Science.gov (United States)

    Gomez, Federico J V; Monasterio, Romina P; Vargas, Verónica Carolina Soto; Silva, María F

    2012-08-01

    The accurate determination of marker chemical species in grape, musts, and wines presents a unique analytical challenge with high impact on diverse areas of knowledge such as health, plant physiology, and economy. Capillary electromigration techniques have emerged as a powerful tool, allowing the separation and identification of highly polar compounds that cannot be easily separated by traditional HPLC methods, providing complementary information and permitting the simultaneous analysis of analytes with different nature in a single run. The main advantage of CE over traditional methods for wine analysis is that in most cases samples require no treatment other than filtration. The purpose of this article is to present a revision on capillary electromigration methods applied to the analysis of wine and its precursors over the last decade. The current state of the art of the topic is evaluated, with special emphasis on the natural compounds that have allowed wine to be considered as a functional food. The most representative revised compounds are phenolic compounds, amino acids, proteins, elemental species, mycotoxins, and organic acids. Finally, a discussion on future trends of the role of capillary electrophoresis in the field of analytical characterization of wines for routine analysis, wine classification, as well as multidisciplinary aspects of the so-called "from soil to glass" chain is presented. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. A method for easily customizable gradient gel electrophoresis.

    Science.gov (United States)

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

    2016-09-15

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

  13. Instrumental development of novel detection and separation methods for capillary electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Garner, Tommy [Iowa State Univ., Ames, IA (United States)

    1993-07-01

    After a general introduction, this thesis is divided into 3 parts: indirect fluorescence detection of sugars separated by capillary zone electrophoresis with visible laser excitation, absorption detection in capillary electrophoresis by fluorescence energy transfer, and increased selectivity for electrochromatography by dynamic ion exchange.

  14. Nonradioactive telomerase activity assay by microchip electrophoresis: privileges to the classical gel electrophoresis assay.

    Science.gov (United States)

    Zhelev, Zhivko; Bakalova, Rumiana; Ewis, Ashraf; Ohba, Hideki; Ishikawa, Mitsuru; Baba, Yoshinobu

    2005-08-01

    The present study accents on the privileges of microchip-based electrophoresis to the conventional gel electrophoresis in separation of telomerase repeat amplification protocol/polymerase chain reaction (PCR) ladder products obtained in telomerase-catalyzed reaction in cancer cells. We try to clarify the interpretation of the results obtained by both electrophoretic procedures and to avoid misinterpretation as a result of PCR-dependent artefacts.

  15. Rapid DNA sequencing by horizontal ultrathin gel electrophoresis.

    OpenAIRE

    Brumley, R L; Smith, L M

    1991-01-01

    A horizontal polyacrylamide gel electrophoresis apparatus has been developed that decreases the time required to separate the DNA fragments produced in enzymatic sequencing reactions. The configuration of this apparatus and the use of circulating coolant directly under the glass plates result in heat exchange that is approximately nine times more efficient than passive thermal transfer methods commonly used. Bubble-free gels as thin as 25 microns can be routinely cast on this device. The appl...

  16. Stacking gels: A method for maximising output for pulsed-field gel electrophoresis

    Directory of Open Access Journals (Sweden)

    Heng See

    2009-01-01

    Full Text Available Pulsed field gel electrophoresis (PFGE, the gold standard of molecular typing methods, has a major disadvantage of an unusually long electrophoretic time. From the original protocol of 6 days, it was modified to 3 days and subsequently to a single day. We describe the procedure of stacking five to six gels one on top of another in order to increase and maximize the output in a shorter time without compromising the resolution and reproducibility. All the variables that affect pulsed field gels during electrophoresis were taken into consideration. We firstly optimized the parameters to be used and secondly determined whether stacking of five to six gels had any effect on the molecular separation during electrophoresis in comparison with a single gel run. DNA preparation, restriction, electrophoresis, staining and gel documentation was carried out based on previously published methods. Gels were analysed using BioNumerics and dice coefficient and unweighted pair group methods were used to generate dendrograms based on 1.5% tolerance values. Identical band profiles and band resolution-separation were seen in the PFGE patterns with single gel and multiple stacking gels. Cluster analysis further strengthened the fact that results from stacking gels were reproducible and comparable with a single gel run. This method of stacking gels saves time and maximizes the output at the same time. The run time for a single gel was about 28 hours, but with six stacked gels the run time was 54 hours compared with 28 x 6 = 168 hours if they were run separately as single gels thus saving time of 67.86%. Beside the big factor of saving time, stacking gels save resources (electricity, reagents, water, chemicals and working time by increasing the sample throughput in a shorter time without compromising on quality of data. But optimization of working parameters is vital depending on the PFGE system used.

  17. Explorative data analysis of two-dimensional electrophoresis gels

    DEFF Research Database (Denmark)

    Schultz, J.; Gottlieb, D.M.; Petersen, Marianne Kjerstine

    2004-01-01

    of gels is presented. First, an approach is demonstrated in which no prior knowledge of the separated proteins is used. Alignment of the gels followed by a simple transformation of data makes it possible to analyze the gels in an automated explorative manner by principal component analysis, to determine......Methods for classification of two-dimensional (2-DE) electrophoresis gels based on multivariate data analysis are demonstrated. Two-dimensional gels of ten wheat varieties are analyzed and it is demonstrated how to classify the wheat varieties in two qualities and a method for initial screening...... if the gels should be further analyzed. A more detailed approach is done by analyzing spot volume lists by principal components analysis and partial least square regression. The use of spot volume data offers a mean to investigate the spot pattern and link the classified protein patterns to distinct spots...

  18. [Determination of glutamic acid in biological material by capillary electrophoresis].

    Science.gov (United States)

    Narezhnaya, E; Krukier, I; Avrutskaya, V; Degtyareva, A; Igumnova, E A

    2015-01-01

    The conditions for the identification and determination of Glutamic acid by capillary zone electrophoresis without their preliminary derivatization have been optimized. The effect of concentration of buffer electrolyte and pH on determination of Glutamic acid has been investigated. It is shown that the 5 Mm borate buffer concentration and a pH 9.15 are optimal. Quantitative determination of glutamic acid has been carried out using a linear dependence between the concentration of the analyte and the area of the peak. The accuracy and reproducibility of the determination are confirmed by the method "introduced - found". Glutamic acid has been determined in the placenta homogenate. The duration of analysis doesn't exceed 30 minutes. The results showed a decrease in the level of glutamic acid in cases of pregnancy complicated by placental insufficiency compared with the physiological, and this fact allows to consider the level of glutamic acid as a possible marker of complicated pregnancy.

  19. 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.

  20. Determination of Betaine in Lycii Cortex by Capillary Electrophoresis

    Science.gov (United States)

    Peng, Xuewei; Liu, Haixing

    2017-12-01

    This paper presents the determination of betaine content in Lycii Cortex by high performance capillary electrophoresis (HPCE) method. The borax solution was chosen as buffer solution, and its concentration was 40 mmol at a constant voltage of 20kV and injecting pressure time of 10s at 14°C. Linearity was kept in the concent ration range of 0.0113∼1.45mg of betaine with correlation coefficient of 0.9. The content of betaine in Lycii Cortex was 61.9 mg/g (RSD = 13.4%) (n = 7). The recovery was in the range of 86.6% - 118.1% (n=4). This method is specific, simple and rapid and accurate, which is suitable for the detection of the content of betaine in Lycii Cortex.

  1. Determination of Betaine in Jujube by Capillary Electrophoresis

    Science.gov (United States)

    Han, Likun; Liu, Haixing; Peng, Xuewei

    2017-12-01

    This paper presents the determination of betaine content in jujube by high performance capillary electrophoresis (HPCE) method. The borax solution was chosen as buffer solution, and its concentration was 40 mmol at a constant voltage of 20kV and injecting pressure time of 10s at 14°C. Linearity was kept in the concent ration range of 0.0113∼1.45mg of betaine with correlation coefficient of 0.9. The content of betaine in jujube was 85.91 mg/g (RSD = 16.6%) (n = 6). The recovery of betaine in jujube sample was in the range of 86.2% - 116.6% (n=3). This method is specific, simple and rapid and accurate, which is suitable for the detection of the content of betaine in jujube.

  2. Capillary sieving electrophoresis and micellar electrokinetic capillary chromatography produce highly correlated separation of tryptic digests

    Science.gov (United States)

    Dickerson, Jane A.; Dovichi, Norman J.

    2011-01-01

    We perform two-dimensional capillary electrophoresis on fluorescently labeled proteins and peptides. Capillary sieving electrophoresis was performed in the first dimension and micellar electrokinetic capillary chromatography was performed in the second. A cellular homogenate was labeled with the fluorogenic reagent FQ and separated using the system. This homogenate generated a pair of ridges; the first had essentially constant migration time in the CSE dimension, while the second had essentially constant migration time in the MEKC dimension. In addition a few spots were scattered through the electropherogram. The same homogenate was digested using trypsin, and then labeled and subjected to the two dimensional separation. In this case, the two ridges observed from the original two-dimensional separation disappeared, and were replaced by a set of spots that fell along the diagonal. Those spots were identified using a local-maximum algorithm and each was fit using a two-dimensional Gaussian surface by an unsupervised nonlinear least squares regression algorithm. The migration times of the tryptic digest components were highly correlated (r = 0.862). When the slowest migrating components were eliminated from the analysis, the correlation coefficient improved to r = 0.956. PMID:20564272

  3. Defining carbohydrate binding of glucan phosphatases via Affinity gel electrophoresis

    DEFF Research Database (Denmark)

    Auger, Kyle; Raththagala, Madushi; Wilkens, Casper

    2016-01-01

    was to determine a technique to measure carbohydrate binding quickly and efficiently. We established a protocol to reproducibly and quantitatively measure the binding of the enzymes to glucans utilizing Affinity Gel Electrophoresis (AGE). The results show that the various glucan phosphatases possess differing...

  4. Gel Electrophoresis and Fluorescamine Methods for the Detection of ...

    African Journals Online (AJOL)

    For the fluorescamine method, clarification was achieved by isoelectric precipitation and precipitation with acid to obtain pH 4.6 and 6% TCA soluble extracts respectively. Non-clarified samples were used for gel electrophoresis. Both methods confirmed that raw milk and milk processed at 85/15s were the most proteolysed, ...

  5. Strain identification in Rhizobium by starch gel electrophoresis of isoenzymes

    DEFF Research Database (Denmark)

    Engvild, Kjeld Christensen; Nielsen, G.

    1985-01-01

    Sonieated extracts of rhizobia, especiaUy Rhizobium leguminosarum from pea and vetch, were run in horizontal starch gel electrophoresis in the cold. The rhizobia were grown on agar on a slime suppressing substrate of tryptone-yeast extract-CaCl2 with small amounts of mannitol, sorbitol...

  6. Analysis of Two-Dimensional Electrophoresis Gel Images

    DEFF Research Database (Denmark)

    Pedersen, Lars

    2002-01-01

    This thesis describes and proposes solutions to some of the currently most important problems in pattern recognition and image analysis of two-dimensional gel electrophoresis (2DGE) images. 2DGE is the leading technique to separate individual proteins in biological samples with many biological...

  7. Pulsed-field gel electrophoresis typing of Staphylococcus aureus isolates

    Science.gov (United States)

    Pulsed-field gel electrophoresis (PFGE) is the most applied and effective genetic typing method for epidemiological studies and investigation of foodborne outbreaks caused by different pathogens, including Staphylococcus aureus. The technique relies on analysis of large DNA fragments generated by th...

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

    African Journals Online (AJOL)

    Denaturing gradient gel electrophoresis (DGGE) is a powerful method used to study structure of bacterial communities, without cultivation, based on the diversity of the genes coding for ribosomal RNA. However, the results are strongly dependent on the respective target region of the used primer systems. Therefore, three ...

  9. Two-dimensional gel electrophoresis analysis of different parts of ...

    African Journals Online (AJOL)

    Two-dimensional gel electrophoresis analysis of different parts of Panax quinquefolius L. root. ... From these results it was concluded that proteomic analysis method was an effective way to identify the different parts of quinquefolius L. root. These findings may contribute to further understanding of the physiological ...

  10. The single-cell gel electrophoresis assay to determine apoptosis ...

    African Journals Online (AJOL)

    When the frequency of appearance of apoptotic cells following was observed over a period of time, there was a significant increase in appearance of apoptosis when using single cell gel electrophoresis assay. The present report demonstrates that the characteristic pattern of apoptotic comets detected by the comet assay ...

  11. Acid-Urea Gel Electrophoresis and Western Blotting of Histones.

    Science.gov (United States)

    Hazzalin, Catherine A; Mahadevan, Louis C

    2017-01-01

    Acid-urea gel electrophoresis offers significant advantages over SDS-PAGE for analysis of post-translational protein modifications, being capable of resolving proteins of similar size but varying in charge. Hence, it can be used to separate protein variants with small charge-altering differences in primary sequence, and is particularly useful in the analysis of histones whose charge variation arises from post-translational modification, such as phosphorylation or acetylation. On acid-urea gels, histones that carry multiple modifications, each with a characteristic charge, are resolved into distinct bands, the so-called "histone ladder." Thus, the extent and distribution of different modification states of histones can be visualized. Here, we describe the analysis of histone H3 by acid-urea gel electrophoresis and western blotting.

  12. Parallel analysis and orthogonal identification of N-glycans with different capillary electrophoresis mechanisms

    International Nuclear Information System (INIS)

    Feng, Hua-tao; Su, Min; Rifai, Farida Nur; Li, Pingjing; Li, Sam F.Y.

    2017-01-01

    The deep involvement of glycans or carbohydrate moieties in biological processes makes glycan patterns an important direction for the clinical and medicine researches. A multiplexing CE mapping method for glycan analysis was developed in this study. By applying different CE separation mechanisms, the potential of combined parallel applications of capillary zone electrophoresis (CZE), micellar electrokinetic chromatography (MEKC) and capillary gel electrophoresis (CGE) for rapid and accurate identification of glycan was investigated. The combination of CZE and MEKC demonstrated enhancing chromatography separation capacity without the compromises of sample pre-treatment and glycan concentration. The separation mechanisms for multiplexing platform were selected based on the orthogonalities of the separation of glycan standards. MEKC method exhibited promising ability for the analysis of small GU value glycans and thus complementing the unavailability of CZE. The method established required only small amount of samples, simple instrument and single fluorescent labelling for sensitive detection. This integrated method can be used to search important glycan patterns appearing in biopharmaceutical products and other glycoproteins with clinical importance. - Highlights: • Cross-validation of analytes in complex samples was done with different CE separation mechanisms. • A simple strategy is used to confirm peak identification and extend capacity of CE separation. • The method uses small amount of sample, simple instrument and single fluorescent labeling. • Selection of mechanisms is based on orthogonalities of GU values of glycan standards. • Micellar electrokinetic chromatography was suitable for analysis of small or highly sialylated glycans.

  13. Parallel analysis and orthogonal identification of N-glycans with different capillary electrophoresis mechanisms

    Energy Technology Data Exchange (ETDEWEB)

    Feng, Hua-tao [Department of Chemistry, National University of Singapore, 3 Science Drive 3, Singapore 117543 (Singapore); NUS Environmental Research Institute, 5A Engineering Drive 1, T-Lab Building, Singapore 117411 (Singapore); Su, Min; Rifai, Farida Nur [Department of Chemistry, National University of Singapore, 3 Science Drive 3, Singapore 117543 (Singapore); Li, Pingjing [NUS Environmental Research Institute, 5A Engineering Drive 1, T-Lab Building, Singapore 117411 (Singapore); Li, Sam F.Y., E-mail: chmlifys@nus.edu.sg [Department of Chemistry, National University of Singapore, 3 Science Drive 3, Singapore 117543 (Singapore); NUS Environmental Research Institute, 5A Engineering Drive 1, T-Lab Building, Singapore 117411 (Singapore)

    2017-02-08

    The deep involvement of glycans or carbohydrate moieties in biological processes makes glycan patterns an important direction for the clinical and medicine researches. A multiplexing CE mapping method for glycan analysis was developed in this study. By applying different CE separation mechanisms, the potential of combined parallel applications of capillary zone electrophoresis (CZE), micellar electrokinetic chromatography (MEKC) and capillary gel electrophoresis (CGE) for rapid and accurate identification of glycan was investigated. The combination of CZE and MEKC demonstrated enhancing chromatography separation capacity without the compromises of sample pre-treatment and glycan concentration. The separation mechanisms for multiplexing platform were selected based on the orthogonalities of the separation of glycan standards. MEKC method exhibited promising ability for the analysis of small GU value glycans and thus complementing the unavailability of CZE. The method established required only small amount of samples, simple instrument and single fluorescent labelling for sensitive detection. This integrated method can be used to search important glycan patterns appearing in biopharmaceutical products and other glycoproteins with clinical importance. - Highlights: • Cross-validation of analytes in complex samples was done with different CE separation mechanisms. • A simple strategy is used to confirm peak identification and extend capacity of CE separation. • The method uses small amount of sample, simple instrument and single fluorescent labeling. • Selection of mechanisms is based on orthogonalities of GU values of glycan standards. • Micellar electrokinetic chromatography was suitable for analysis of small or highly sialylated glycans.

  14. Indirect fluorescence detection of native amino acids in capillary zone electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Kuhr, W.G.; Yeung, E.S.

    1988-09-01

    Amino acids are but one of several important classes of small chemical compounds in biological chemistry that have an inherent lack of analytically useful physical properties. Amino acids, peptides, fatty acids, sugars, many mono-, di-, and tricarboxylic acids, and phosphorylated intermediates in glycolysis and metabolism show little, if any, UV or visible absorption, fluorescence, or electrochemical activity. As the emphasis of biochemical research shifts to smaller samples where, for example, picomolar quantities of amino acids are analyzed in gas phase protein sequencing or in microliter samples of the extracellular fluid of the mammalian brain, the analytical problem becomes even more challenging due to the small volume of sample available for analysis. In this work, laser-induced fluorescence spectroscopy is performed on-column to detect the bands separated with capillary zone electrophoresis (CZE). CZE is an instrumental form of zone electrophoresis where chemical species are separated purely on the basis of their electrophoretic mobility, since no supporting gel is utilized. Both anions and cations can be separated in the same run because of the large electroosmotic flow generated in small diameter capillaries. This technique has already been used successfully in the rapid, efficient separation of dansyl-amino acids.

  15. Rapid capillary coating by epoxy-poly-(dimethylacrylamide): Performance in capillary zone electrophoresis of protein and polystyrene carboxylate

    Czech Academy of Sciences Publication Activity Database

    Chiari, M.; Cretich, M.; Šťastná, Miroslava; Radko, S. P.; Chrambach, A.

    2001-01-01

    Roč. 22, č. 4 (2001), s. 656-659 ISSN 0173-0835 Institutional research plan: CEZ:AV0Z4031919 Keywords : capillary coating * capillary zone electrophoresis * proteins Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 4.282, year: 2001

  16. High lane density slab-gel electrophoresis using micromachined instrumentation.

    Science.gov (United States)

    Papautsky, I; Mohanty, S; Weiss, R; Frazier, A B

    2001-10-01

    In this paper, micromachined pipette arrays (MPAs) and microcombs were studied as a means of enabling high lane density gel electrophoresis. The MPA provide a miniaturized format to interface sub-microliter volumes of samples between macroscale sample preparation formats and microscale biochemical analysis systems. The microcombs provide a means of creating sample loading wells in the gel material on the same center-to-center spacing as the MPAs. Together, the two micromachined instruments provide an alternative to current combs and pipetting technologies used for creating sample loading wells and sample delivery in gel electrophoresis systems. Using three designs for the microcomb-MPA pair, center-to-center spacings of 1.0 mm, 500 microm, and 250 microm are studied. The results demonstrate an approximate 10-fold increase in lane density and a 10-fold reduction in sample size from 5 microL to 500 pL. As a result, the number of theoretical plates has increased 2.5-fold, while system resolution has increased 1.5-fold over the conventional agarose gel systems. An examination of changes in resolution across the width of individual separation lanes in both systems revealed dependence in the case of the conventional gels and no dependence for the gels loaded with the micromachined instrumentation.

  17. Determination of dioxopromethazine hydrochloride by capillary electrophoresis with electrochemiluminescence detection

    International Nuclear Information System (INIS)

    Li Yunhui; Wang Chunyan; Sun Jinying; Zhou Yongchang; You Tianyan; Wang Erkang; Fung Yingsing

    2005-01-01

    The paper presents a rapid method for the determination of dioxopromethazine hydrochloride (DPZ), an antihistamine drug, by the capillary electrophoresis with electrochemiluminescene detection (CE-ECL) using tris(2,2'-bipyridyl)ruthenium(II) (Ru(bpy) 3 2+ ) reagent. This CE-ECL detection method has high sensitivity, good selectivity and reproducibility for DPZ analysis. Under the optimized conditions: separation capillary, 38 cm length (25 μm i.d.); sample injection, 10 s at 8 kV; separation voltage, 12.5 kV; running buffer, 20 mmol L -1 sodium phosphate of pH 6.0; detection potential, 1.15 V; 50 mmol L -1 of phosphate buffer (pH 7.14) containing 5 mmol L -1 of Ru(bpy) 3 2+ in ECL detection cell, the detection limit of DPZ was 0.05 μmol L -1 (S/N = 3). The linear range extended from 5 to 100 μmol L -1 . The linear curve obtained was Y = 181.62 + 9.28X with a correlation coefficient of 0.9970. The relative standard deviations of the ECL intensity and the migration time for six continuous injections of 5 μmol L -1 DPZ were 3.7% and 0.92%, respectively. The CE-ECL method was applied to analyze DPZ in real samples including tablets, rat serum and human urine, and satisfactory results were obtained without interference from samples matrix. The CE-ECL technique was proved to be a potential method for the detection of DPZ in clinic analysis

  18. Images of gel electrophoresis - RGP caps | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available List Contact us RGP caps Images of gel electrophoresis Data detail Data name Images of gel electrophoresis D...OI 10.18908/lsdba.nbdc00318-05-002 Description of data contents Detailed information and images of gel electrophoresis... of each marker. Data file File name: rgp_caps_electrophoresis_image.zip File URL: ftp://ftp.biosc...iencedbc.jp/archive/rgp-caps/LATEST/rgp_caps_electrophoresis_image.zip File size:... 28.7 MB Simple search URL - Data acquisition method Gel electrophoresis Data analysis method STS markers :

  19. Blood grouping based on PCR methods and agarose gel electrophoresis.

    Science.gov (United States)

    Sell, Ana Maria; Visentainer, Jeane Eliete Laguila

    2015-01-01

    The study of erythrocyte antigens continues to be an intense field of research, particularly after the development of molecular testing methods. More than 300 specificities have been described by the International Society for Blood Transfusion as belonging to 33 blood group systems. The polymerase chain reaction (PCR) is a central tool for red blood cells (RBC) genotyping. PCR and agarose gel electrophoresis are low cost, easy, and versatile in vitro methods for amplifying defined target DNA (RBC polymorphic region). Multiplex-PCR, AS-PCR (Specific Allele Polymerase Chain Reaction), and RFLP-PCR (Restriction Fragment Length Polymorphism-Polymerase Chain Reaction) techniques are usually to identify RBC polymorphisms. Furthermore, it is an easy methodology to implement. This chapter describes the PCR methodology and agarose gel electrophoresis to identify the polymorphisms of the Kell, Duffy, Kidd, and MNS blood group systems.

  20. 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.

  1. Interaction of albumins and heparinoids investigated by affinity capillary electrophoresis and free flow electrophoresis.

    Science.gov (United States)

    Mozafari, Mona; El Deeb, Sami; Krull, Friederike; Wildgruber, Robert; Weber, Gerhard; Reiter, Christian G; Wätzig, Hermann

    2018-02-01

    A fast and precise affinity capillary electrophoresis (ACE) method has been applied to investigate the interactions between two serum albumins (HSA and BSA) and heparinoids. Furthermore, different free flow electrophoresis methods were developed to separate the species which appears owing to interaction of albumins with pentosan polysulfate sodium (PPS) under different experimental conditions. For ACE experiments, the normalized mobility ratios (∆R/R f ), which provided information about the binding strength and the overall charge of the protein-ligand complex, were used to evaluate the binding affinities. ACE experiments were performed at two different temperatures (23 and 37°C). Both BSA and HSA interact more strongly with PPS than with unfractionated and low molecular weight heparins. For PPS, the interactions can already be observed at low mg/L concentrations (3 mg/L), and saturation is already obtained at approximately 20 mg/L. Unfractionated heparin showed almost no interactions with BSA at 23°C, but weak interactions at 37°C at higher heparin concentrations. The additional signals also appeared at higher concentrations at 37°C. Nevertheless, in most cases the binding data were similar at both temperatures. Furthermore, HSA showed a characteristic splitting in two peaks especially after interacting with PPS, which is probably attributable to the formation of two species or conformational change of HSA after interacting with PPS. The free flow electrophoresis methods have confirmed and completed the ACE experiments. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  2. Integration of capillary electrophoresis with gold nanoparticle-based colorimetry.

    Science.gov (United States)

    Li, Tong; Wu, Zhenglong; Qin, Weidong

    2017-12-01

    A method integrating capillary electrophoresis (CE) and gold nanoparticle aggregation-based colorimetry (AuNP-ABC) was described. By using a dual-sheath interface, the running buffer was isolated from the colorimetric reaction solution so that CE and AuNP-ABC would not interfere with each other. The proof-of-concept was validated by assay of polyamidoamine (PAMAM) dendrimers that were fortified in human urine samples. The factors influencing the CE-AuNP-ABC performances were investigated and optimized. Under the optimal conditions, the dendrimers were separated within 8 min, with detection limits of 0.5, 1.2 and 2.6 μg mL -1 for PAMAM G1.0, G2.0 and G3.0, respectively. The sensitivity of CE-AuNP-ABC was comparable to or even better than those of liquid chromatography-fluorimetry and liquid chromatography-mass spectrometry. The results suggested that the proposed strategy can be applied to facile and quick determination of analytes of similar properties in complex matrices. Copyright © 2017 Elsevier B.V. All rights reserved.

  3. Quantification of carbamylated albumin in serum based on capillary electrophoresis.

    Science.gov (United States)

    Delanghe, Sigurd; Moerman, Alena; Pletinck, Anneleen; Schepers, Eva; Glorieux, Griet; Van Biesen, Wim; Delanghe, Joris R; Speeckaert, Marijn M

    2017-09-01

    Protein carbamylation, a nonenzymatic posttranslational modification promoted during uremia, is linked to a poor prognosis. In the present study, carbamylation of serum albumin was assayed using the symmetry factor on a capillary electrophoresis instrument (Helena V8). The symmetry factor has been defined as the distance from the center line of the peak to the back slope, divided by the distance from the center line of the peak to the front slope, with all measurements made at 10% of the maximum peak height. Serum albumin, creatinine, and urea concentrations were assayed using routine methods, whereas uremic toxins were determined using HPLC. In vitro carbamylation induced a marked albumin peak asymmetry. Reference values for the albumin symmetry factor were 0.69-0.92. In kidney patients, albumin peak asymmetry corresponded to the chronic kidney disease stage (p < 0.0001). The symmetry factor correlated well with serum urea (r = -0.5595, p < 0.0001) and creatinine (r = -0.5986, p < 0.0001) concentrations. Several protein-bound uremic toxins showed a significant negative correlation with the symmetry factor. Morphology of the albumin fraction was not affected by presence of glycated albumin and protein-bound antibiotics. In conclusion, the presented method provides a simple, practical way for monitoring protein carbamylation. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. Capillary electrophoresis methods for microRNAs assays: A review

    Energy Technology Data Exchange (ETDEWEB)

    Ban, Eunmi; Song, Eun Joo, E-mail: ejsong@kist.re.kr

    2014-12-10

    Highlights: • A review of CE analysis of miRNAs. • Summary of developments and applications of CE systems in miRNA studies. • Applications and development of microchip-based CE for rapid analysis of miRNA. - Abstract: MicroRNAs (miRNAs) are short noncoding RNAs that conduct important roles in many cellular processes such as development, proliferation, differentiation, and apoptosis. In particular, circulating miRNAs have been proposed as biomarkers for cancer, diabetes, cardiovascular disease, and other illnesses. Therefore, determination of miRNA expression levels in various biofluids is important for the investigation of biological processes in health and disease and for discovering their potential as new biomarkers and drug targets. Capillary electrophoresis (CE) is emerging as a useful analytical tool for analyzing miRNA because of its simple sample preparation steps and efficient resolution of a diverse size range of compounds. In particular, CE with laser-induced fluorescence detection is a promising and relatively rapidly developing tool with the potential to provide high sensitivity and specificity in the analysis of miRNAs. This paper covers a short overview of the recent developments and applications of CE systems in miRNA studies in biological and biomedical areas.

  5. Demonstrate use of capillary electrophoresis low level transient of anions

    International Nuclear Information System (INIS)

    Moum, Kari-Lye; Solheim, Torill; McElrath, Joel; Frattini, Paul

    2012-09-01

    Capillary Electrophoresis (CE) is a well-known analytical method capable of rapid detection of very low concentration of cations and anionic species such as chloride, sulfate and nitrate. These anions are of crucial importance in reducing the potential of stainless steel components to undergo stress corrosion cracking. Currently, Nuclear Power Plants (NPPs) use Ion Chromatography (IC) as the analytical technique to achieve the required detection levels of ionic species. At the Halden Reactor Project (HRP) IC was replaced by CE in 1996, and since then HRP has gained nearly 20 years of operational experience. During the last 15 years, EPRI has done research on the CE technique and has achieved extensive experience in this area. EPRI has demonstrated detection levels at ppt and sub-ppb levels. This paper presents the ability of the CE technique to follow low level transients of anions in Boiling Water Reactor (BWR) coolant. A transient caused by approx. 10 ppb chloride and sulfate was simulated in an experimental circuit simulating BWR conditions. A series of grab samples were taken and analysed using HRPs CE (Agilent G1600). (authors)

  6. Trace analysis of organic ions in ice samples by capillary electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Huber, T. [Bern Univ. (Switzerland); Schwikowski, M.; Gaeggeler, H.W. [Paul Scherrer Inst. (PSI), Villigen (Switzerland)

    1997-09-01

    Capillary electrophoresis was tested as a new analytical method for ice samples. Comparisons to ion chromatography were made concerning accuracy, detection limits, reproducibility, necessary sample volume and time consumption. (author) 1 fig., 3 refs.

  7. ANALYSIS OF THE ENANTIOMERS OF CHIRAL PESTICIDES AND OTHER POLLUTANTS IN ENVIRONMENTAL SAMPLES BY CAPILLARY ELECTROPHORESIS

    Science.gov (United States)

    The generic method described here involves typical capillary electrophoresis (CE) techniques, with the addition of cyclodextrin chiral selectors to the electrolyte for enantiomer separation and also, in the case of neutral analytes, the further addition of a micelle forming comp...

  8. Trace analysis of organic ions in ice samples by capillary electrophoresis

    International Nuclear Information System (INIS)

    Huber, T.; Schwikowski, M.; Gaeggeler, H.W.

    1997-01-01

    Capillary electrophoresis was tested as a new analytical method for ice samples. Comparisons to ion chromatography were made concerning accuracy, detection limits, reproducibility, necessary sample volume and time consumption. (author) 1 fig., 3 refs

  9. 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...

  10. Role of capillary electrophoresis in the fight against doping in sports.

    Science.gov (United States)

    Harrison, Christopher R

    2013-08-06

    At present the role of capillary electrophoresis in the detection of doping agents in athletes is, for the most part, nonexistent. More traditional techniques, namely gas and liquid chromatography with mass spectrometric detection, remain the gold standard of antidoping tests. This Feature will investigate the in-roads that capillary electrophoresis has made, the limitations that the technique suffers from, and where the technique may grow into being a key tool for antidoping analysis.

  11. VIII All-Russian symposium on molecular liquid chromatography and capillary electrophoresis. Program. Summary of reports

    International Nuclear Information System (INIS)

    2001-01-01

    Program and summary of reports of the VIII All-Russian symposium on molecular liquid chromatography and capillary electrophoresis are performed. The meeting took place 15-19 October, 2001 in Moscow. Many problems of liquid and ion exchange chromatography, capillary electrophoresis, thin-layer chromatography have been discussed extensively. Reports covering properties of sorbents and devices for chromatography are incorporated in the collection [ru

  12. Sample Stacking in capillary zone electrophoresis : Principles, advantages and limitations

    NARCIS (Netherlands)

    Beckers, J.L.; Bocek, P.

    2000-01-01

    The principles of stacking procedures are described and their properties are discussed, including the fundamentals of the behavior of zone boundaries and the consequences of the self-correcting properties of boundaries in moving boundary electrophoresis, isotachophoresis, and zone electrophoresis.

  13. Improving precision in gel electrophoresis by stepwisely decreasing variance components.

    Science.gov (United States)

    Schröder, Simone; Brandmüller, Asita; Deng, Xi; Ahmed, Aftab; Wätzig, Hermann

    2009-10-15

    Many methods have been developed in order to increase selectivity and sensitivity in proteome research. However, gel electrophoresis (GE) which is one of the major techniques in this area, is still known for its often unsatisfactory precision. Percental relative standard deviations (RSD%) up to 60% have been reported. In this case the improvement of precision and sensitivity is absolutely essential, particularly for the quality control of biopharmaceuticals. Our work reflects the remarkable and completely irregular changes of the background signal from gel to gel. This irregularity was identified as one of the governing error sources. These background changes can be strongly reduced by using a signal detection in the near-infrared (NIR) range. This particular detection method provides the most sensitive approach for conventional CCB (Colloidal Coomassie Blue) stained gels, which is reflected in a total error of just 5% (RSD%). In order to further investigate variance components in GE, an experimental Plackett-Burman screening design was performed. The influence of seven potential factors on the precision was investigated using 10 proteins with different properties analyzed by NIR detection. The results emphasized the individuality of the proteins. Completely different factors were identified to be significant for each protein. However, out of seven investigated parameters, just four showed a significant effect on some proteins, namely the parameters of: destaining time, staining temperature, changes of detergent additives (SDS and LDS) in the sample buffer, and the age of the gels. As a result, precision can only be improved individually for each protein or protein classes. Further understanding of the unique properties of proteins should enable us to improve the precision in gel electrophoresis.

  14. Two-dimensional polyacrylamide gel electrophoresis of intracellular proteins

    International Nuclear Information System (INIS)

    Ojima, N.; Sakamoto, T.; Yamashita, M.

    1996-01-01

    Since two-dimensional electrophoresis was established by O'Farrell for analysis of intracellular proteins of Escherichia coli, it has been applied to separation of proteins of animal cells and tissues, and especially to identification of stress proteins. Using this technique, proteins are separated by isoelectric focusing containing 8 m urea in the first dimension and by SDS-PAGE in the second dimension. The gels are stained with Coomassie Blue R-250 dye, followed by silver staining. In the case of radio-labeled proteins, the gels are dried and then autoradiographed. In order to identify a specific protein separated by two-dimensional electrophoresis, a technique determining the N-terminal amino acid sequence of the protein has been developed recently. After the proteins in the gel were electrotransferred to a polyvinylidene difluoride membrane, the membrane was stained for protein with Commassie Blue and a stained membrane fragment was applied to a protein sequencer. Our recent studies demonstrated that fish cells newly synthesized various proteins in response to heat shock, cold nd osmotic stresses. For example, when cellular proteins extracted from cold-treated rainbow trout cells were subjected to two-dimensional gel electrophoresis, the 70 kDa protein was found to be synthesized during the cold-treatment. N-Terminal sequence analysis showed that the cold-inducible protein was a homolog of mammalian valosin-containing protein and yeast cell division cycle gene product CDC48p. Furthermore, the sequence data were useful for preparing PCR primers and a rabbit antibody against a synthetic peptide to analyze a role for the protein in the function of trout cells and mechanisms for regulation

  15. Choice of capillary electrophoresis systems for the impurity profiling of drugs

    NARCIS (Netherlands)

    Hilhorst, M.J; Somsen, G.W; de Jong, G.J.

    In order to develop a strategy for the impurity profiling of drugs, the possibilities of some capillary electrophoresis systems were investigated. A mixture containing a drug and some of its possible impurities has been used as a model problem. The test compounds were investigated by capillary zone

  16. Analytical characterization of heparin by capillary zone electrophoresis with conductivity detection and polymeric buffer additives.

    Science.gov (United States)

    Mikus, Peter; Valásková, Iva; Havránek, Emil

    2004-11-15

    A capillary zone electrophoresis (CZE) method for the analytical characterization of intact (high-molecular-weight) heparin was developed. For the first time, a hydrodynamically closed CZE separation system with conductivity detector was used for the separation, detection and quantitation of this highly sulfated, linear polysaccharide. Glycine (25mM) adjusted to pH 9.0 by bis-Tris-propane served as the running electrolyte system. Polymeric additives, polyvinylpyrrolidone (PVP), dextran (DEX), were used to improve the separation selectivity as they strongly retarded the heparin macromolecule while they did not practically influence comigrating inorganic anions. The proposed electrophoretic method was successfully validated. It was convenient for the sensitive, simple, rapid and reproducible assay of heparin in raw materials and isotonic saline. Here, the use of the conductivity detector was advantageous as it allowed heparin to be analyzed without a sample pretreatment. The CZE method should be an alternative to the pharmacopoeial conventional gel electrophoresis having used in the quality control of heparin so far. In addition, it should be convenient to quantitative estimation of heparin present in a preparation used, e.g., as the chiral selector in CE separations.

  17. Exploring chip-capillary electrophoresis-laser-induced fluorescence field-deployable platform flexibility: Separations of fluorescent dyes by chip-based non-aqueous capillary electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Nuchtavorn, N.; Smejkal, Petr; Breadmore, M. C.; Guijt, R. M.; Doble, P.; Bek, F.; Foret, František; Suntornsuk, L.; Macka, M.

    2013-01-01

    Roč. 1286, APR (2013), s. 216-221 ISSN 0021-9673 R&D Projects: GA ČR(CZ) GBP206/12/G014 Institutional support: RVO:68081715 Keywords : microfluidic chip CE * capillary electrophoresis * NACE * LIF detection Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 4.258, year: 2013

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

    Science.gov (United States)

    Stellwagen, Nancy C.

    2009-01-01

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

  19. Applications of on-line weak affinity interactions in free solution capillary electrophoresis

    DEFF Research Database (Denmark)

    Heegaard, Niels H H; Nissen, Mogens H; Chen, David D Y

    2002-01-01

    The impressive selectivity offered by capillary electrophoresis can in some cases be further increased when ligands or additives that engage in weak affinity interactions with one or more of the separated analytes are added to the electrophoresis buffer. This on-line affinity capillary...... electrophoresis approach is feasible when the migration of complexed molecules is different from the migration of free molecules and when separation conditions are nondenaturing. In this review, we focus on applying weak interactions as tools to enhance the separation of closely related molecules, e.g., drug...... enantiomers and on using capillary electrophoresis to characterize such interactions quantitatively. We describe the equations for binding isotherms, illustrate how selectivity can be manipulated by varying the additive concentrations, and show how the methods may be used to estimate binding constants. On...

  20. Seed Biology of Medicinal Plants (IX) : The Relationship of Corydalis Species Derived by Gel Electrophoresis

    OpenAIRE

    米田, 該典; 加賀, 順二; 那須, 正夫; KAISUKE, YONEDA; JUNJI, KAGA; MASAO, NASU; 大阪大学薬学部; 大阪大学薬学部; 大阪大学薬学部; Faculty of Pharmaceutical Sciences, Osaka University; Faculty of Pharmaceutical Sciences, Osaka University; Faculty of Pharmaceutical Sciences, Osaka University

    1987-01-01

    The saline soluble protein fraction of seeds of the Corydalis species (Papaveraceae) in Japan was examined by polyacrylamide gel electrophoresis and SDS-polyacrylamide gel electrophoresis. The esterase zymogram suggested that C. pallida, C. pallida var. tenuis, C. heterocarpa var. japonica and C. speciosa, having yellow flowers and no tuber, are closely related to each other. Electrophoresis and SDS-electrophoresis patterns also coincided with the result of the esterase zymogram. They also su...

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

    Science.gov (United States)

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

    2016-08-01

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

  2. Mutation screening of the TP53 gene by temporal temperature gradient gel electrophoresis.

    Science.gov (United States)

    Sørlie, Therese; Johnsen, Hilde; Vu, Phuong; Lind, Guro Elisabeth; Lothe, Ragnhild; Børresen-Dale, Anne-Lise

    2005-01-01

    A protocol for detection of mutations in the TP53 gene using temporal temperature gradient gel electrophoresis (TTGE) is described. TTGE is a mutation detection technique that separates DNA fragments differing by single base pairs according to their melting properties in a denaturing gel. It is based on constant denaturing conditions in the gel combined with a temperature gradient during the electrophoretic run. This method combines some of the advantages of the related techniques denaturing gradient gel electrophoresis (DGGE) and constant denaturant gel electrophoresis (CDGE) and eliminates some of the problems. The result is a rapid and sensitive screening technique that is robust and easily set up in smaller laboratory environments.

  3. Fast separation of enantiomers by capillary electrophoresis using a combination of two capillaries with different internal diameters.

    Science.gov (United States)

    Šebestová, Andrea; Petr, Jan

    2017-12-01

    The combination of capillaries with different internal diameters was used to accelerate the separation of enantiomers in capillary electrophoresis. Separation of R,S-1,1'-binaphthalene-2,2'-diyl hydrogen phosphate using isopropyl derivative of cyclofructan 6 was studied as a model system. The best separation conditions included 500 mM sodium borate pH 9.5 with 60 mM concentration of the chiral selector. Separation lasted approx. 1.5 min using the combination of 50 and 100 μm id capillaries of 9.7 cm and 22.9 cm, respectively. It allowed approx. 12-fold acceleration in comparison to the traditional long-end separation mainly due to the higher electroosmotic flow generated in the connected capillaries. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. Fluid mechanics of electroosmotic flow and its effect on band broadening in capillary electrophoresis.

    Science.gov (United States)

    Ghosal, Sandip

    2004-01-01

    Electroosmotic flow (EOF) usually accompanies electrophoretic migration of charged species in capillary electrophoresis unless special precautions are taken to suppress it. The presence of the EOF provides certain advantages in separations. It is an alternative to mechanical pumps, which are inefficient and difficult to build at small scales, for transporting reagents and analytes on microfluidic chips. The downside is that any imperfection that distorts the EOF profile reduces the separation efficiency. In this paper, the basic facts about EOF are reviewed from the perspective of fluid mechanics and its effect on separations in free solution capillary zone electrophoresis is discussed in the light of recent advances.

  5. Design and operation of a portable scanner for high performance microchip capillary array electrophoresis.

    Science.gov (United States)

    Scherer, James R; Liu, Peng; Mathies, Richard A

    2010-11-01

    We have developed a compact, laser-induced fluorescence detection scanner, the multichannel capillary array electrophoresis portable scanner (McCAEPs) as a platform for electrophoretic detection and control of high-throughput, integrated microfluidic devices for genetic and other analyses. The instrument contains a confocal optical system with a rotary objective for detecting four different fluorescence signals, a pneumatic system consisting of two pressure/vacuum pumps and 28 individual addressable solenoid valves for control of on-chip microvalves and micropumps, four Polymerase Chain Reaction (PCR) temperature control systems, and four high voltage power supplies for electrophoresis. The detection limit of the instrument is ~20 pM for on-chip capillary electrophoresis of fluorescein dyes. To demonstrate the system performance for forensic short tandem repeat (STR) analysis, two experiments were conducted: (i) electrophoretic separation and detection of STR samples on a 96-lane microfabricated capillary array electrophoresis microchip. Fully resolved PowerPlex(®) 16 STR profiles amplified from 1 ng of 9947A female standard DNA were successfully obtained; (ii) nine-plex STR amplification, sample injection, separation, and fluorescence detection of 100-copy 9948 male standard DNA in a single integrated PCR- capillary electrophoresis microchip. These results demonstrate that the McCAEPs can be used as a versatile control and detection instrument that operates integrated microfluidic devices for high-performance forensic human identification.

  6. Recent developments and applications of capillary and microchip electrophoresis in proteomic and peptidomic analyses

    Czech Academy of Sciences Publication Activity Database

    Štěpánová, Sille; Kašička, Václav

    2016-01-01

    Roč. 39, č. 1 (2016), s. 198-211 ISSN 1615-9306 R&D Projects: GA ČR(CZ) GA13-17224S; GA ČR(CZ) GA15-01948S Institutional support: RVO:61388963 Keywords : capillary electrophoresis * mass spectrometry * microchip electrophoresis * peptidomics * proteomics Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 2.557, year: 2016

  7. 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.

  8. Colloid molecular weight estimation by gel chromatography/acrylamide gel electrophoresis

    International Nuclear Information System (INIS)

    Liberatore, F.A.; Dearborn, C.; Nigam, S.; Poon, C.; Camin, L.; Liteplo, M.

    1984-01-01

    Size or molecular weight (MW) estimation of radiolabeled collides in aqueous solutions has long been a problem. The authors have prepared several minimicroaggregated albumin colloids (mμAA) by heat denaturation of stannous-containing HSA solutions at pH 7.0, 7.5, and 8.5). The resulting colloids were labeled with Tc-99m and compared with Au-198 colloid and Tc-99m-antimony sulfide colloid (Tc-99m-Sb/sub 2/S3) by gel chromatography and gel electrophoresis. Tc-99mm-mμAA aggregated at pH 7.0 and the Au-198 colloid appeared in the external void volume of a BioRad A5.0 agarose column indicating an apparent MW of > 5 x 10/sup 6/ daltons. The pH7.5 Tc-99m-mμAA, migrated within the filtration range of the column as did a small fraction of Tc-99m-Sb/sub 2/S/sub 3/, suggesting that the MW is between 6 x 10/sup 4/ - 5 x 10/sup 6/ daltons. The Tc-99m-mμAA, aggregated at pH 8.5, had an apparent MW on gel filtration similar to that of untreated albumin, MW 6.6 x 10-/sup 4/ daltons. The mobilities of the colloids, on acrylamide disc gel electrophoresis, were consistent with the results on gel chromatography. The largest colloids, Au-198 colloid and pH 7.0 Tc-99m-mμAA, barely entered the separating gel; intermediate sized colloids, a small fraction of Tc-99m-Sb/sub 2/S/sub 3/ and pH 7.5 Tc-99m-mμAA migrated farther into the separating gel; while pH 8.5 Tc-99m-mμAA had mobility approaching that of untreated albumin. Lymphoscintigraphy studies using these colloids in animals showed the predicted, particle size-related differences in migration and clearance. The authors conclude that gel chromatography and gel electrophoresis are useful methods for estimating the apparent size of the colloidal particles

  9. Stacking by electroinjection with discontinuous buffers in capillary zone electrophoresis.

    Science.gov (United States)

    Shihabi, Zak K

    2002-08-01

    The work presented here demonstrates that electroinjection can be performed using discontinuous buffers, which can result in better stacking than that obtained by hydrodynamic injection. The sample can be concentrated at the tip of the capillary leaving practically the whole capillary for sample separation. This results in several advantages, such as better sample concentration, higher plate number and shorter time of stacking. However, sample introduction by electromigration is suited for samples free or low in salt content. Samples, which are high in salt content, are better introduced by the hydrodynamic injection for stacking by the discontinuous buffers. Different simple methods to introduce the discontinuity in the buffer for electroinjection are discussed.

  10. Mutation screening of the TP53 gene by temporal temperature gel electrophoresis (TTGE).

    Science.gov (United States)

    Sørlie, Therese; Johnsen, Hilde; Vu, Phuong; Lind, Guro Elisabeth; Lothe, Ragnhild; Børresen-Dale, Anne-Lise

    2014-01-01

    A protocol for detection of mutations in the TP53 gene using temporal temperature gradient electrophoresis (TTGE) is described. TTGE is a mutation detection technique that separates DNA fragments differing by single base pairs according to their melting properties in a denaturing gel. It is based on constant denaturing conditions in the gel combined with a temperature gradient during the electrophoretic run. This method combines some of the advantages of the related techniques, denaturing gradient gel electrophoresis and constant denaturant gel electrophoresis, and eliminates some of the problems. The result is a rapid and sensitive screening technique which is robust and easily set up in smaller laboratory environments.

  11. Capillary Electrophoresis Method Development : Web-based self-paced training-on-demand

    NARCIS (Netherlands)

    Sänger - van de Griend, Cari

    2015-01-01

    If you use capillary electrophoresis (CE) in your work and want a better understanding of the technique, or want to start with CE method development and want to be well prepared, this course is for you. The course is designed for analytical scientists and technicians who use CE in their regular job,

  12. Recent advances in combination of capillary electrophoresis with mass spectrometry: Methodology and theory

    Czech Academy of Sciences Publication Activity Database

    Klepárník, Karel

    2015-01-01

    Roč. 36, č. 1 (2015), s. 159-179 ISSN 0173-0835 R&D Projects: GA ČR(CZ) GA14-28254S Institutional support: RVO:68081715 Keywords : capillary electrophoresis * electrospray * mass spectrometry * Microfluidic devices Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 2.482, year: 2015

  13. Transient isotachophoresis in carrier ampholyte-based capillary electrophoresis for protein analysis

    Czech Academy of Sciences Publication Activity Database

    Busnel, J. M.; Descroix, S.; Godfrin, D.; Hennion, M. C.; Kašička, Václav; Peltre, G.

    2006-01-01

    Roč. 27, č. 18 (2006), s. 3591-3598 ISSN 0173-0835 Institutional research plan: CEZ:AV0Z40550506 Keywords : carrier ampholyte-based capillary electrophoresis * transient isotachophoresis * proteins Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 4.101, year: 2006

  14. Rapid determination of meldonium in urine samples by capillary electrophoresis with capacitively coupled contactless conductivity detection

    Czech Academy of Sciences Publication Activity Database

    Šlampová, Andrea; Kubáň, Pavel

    2016-01-01

    Roč. 1468, OCT (2016), s. 236-240 ISSN 0021-9673 R&D Projects: GA ČR(CZ) GA16-09135S Institutional support: RVO:68081715 Keywords : meldonium * capillary electrophoresis * urine Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.981, year: 2016

  15. Self-aligning subatmospheric hybrid liquid junction electrospray interface for capillary electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Křenková, Jana; Klepárník, Karel; Grym, Jakub; Luksch, Jaroslav; Foret, František

    2016-01-01

    Roč. 37, č. 3 (2016), s. 414-417 ISSN 0173-0835 R&D Projects: GA ČR(CZ) GBP206/12/G014 Institutional support: RVO:68081715 Keywords : capillary electrophoresis * electrospray interfacing * microfabrication Subject RIV: CB - Analytical Chemistry , Separation Impact factor: 2.744, year: 2016

  16. Feasibility of nonvolatile buffers in capillary electrophoresis-electrospray ionization-mass spectrometry of proteins

    NARCIS (Netherlands)

    Eriksson, Jonas H.C.; Mol, Roelof; Somsen, Govert W.; Hinrichs, Wouter L.J.; Frijlink, Henderik W.; de Jong, Gerhardus J.

    2004-01-01

    The combination of capillary electrophoresis (CE) and electrospray ionization-mass spectrometry (ESI-MS) via a triaxial interface was studied as a potential means for the characterization of intact proteins. To evaluate the possibility to use a nonvolatile electrolyte for CE, the effect of sodium

  17. Injections from sub-μL sample volumes in commercial capillary electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Šlampová, Andrea; Kubáň, Pavel

    2017-01-01

    Roč. 1497, MAY (2017), s. 164-171 ISSN 0021-9673 R&D Projects: GA ČR(CZ) GA16-09135S Institutional support: RVO:68081715 Keywords : capillary electrophoresis * microinjections * contactless conductivity detection Subject RIV: CB - Analytical Chemistry, Separation OBOR OECD: Analytical chemistry Impact factor: 3.981, year: 2016

  18. Direct measurement of lithium in whole blood using microchip capillary electrophoresis with integrated conductivity detection

    NARCIS (Netherlands)

    Vrouwe, E.X.; Lüttge, Regina; van den Berg, Albert

    2004-01-01

    The direct measurement of lithium in whole blood is described. Using microchip capillary electrophoresis (CE) with defined sample loading and applying the principles of column coupling, alkali metals were determined in a drop of whole blood. Blood collected from a finger stick was mixed with

  19. Rapid determination of meldonium in urine samples by capillary electrophoresis with capacitively coupled contactless conductivity detection

    Czech Academy of Sciences Publication Activity Database

    Šlampová, Andrea; Kubáň, Pavel

    2016-01-01

    Roč. 1468, OCT (2016), s. 236-240 ISSN 0021-9673 R&D Projects: GA ČR(CZ) GA16-09135S Institutional support: RVO:68081715 Keywords : mel donium * capillary electrophoresis * urine Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.981, year: 2016

  20. Affinity Capillary Electrophoresis – A Powerful Tool to Investigate Biomolecular Interactions

    Czech Academy of Sciences Publication Activity Database

    Kašička, Václav

    2017-01-01

    Roč. 30, č. 5 (2017), s. 248 ISSN 1471-6577 Institutional support: RVO:61388963 Keywords : capillary affinity electrophoresis * biomolecular interactions * binding constants Subject RIV: CB - Analytical Chemistry, Separation OBOR OECD: Analytical chemistry Impact factor: 0.663, year: 2016

  1. SIMULTANEOUS DTERMINATION OF CHROMATE AND AROMATIC HYDROCARBONS IN ENVIRONMENTAL SAMPLES BY CAPILLARY ELECTROPHORESIS

    Science.gov (United States)

    An analytical method was developed to determine simultaneously, the inorganic anion CrO2-4, and organic aromatic compounds including benzoate, 2-Cl-benzoate, phenol, m-cresol and o-/p-cresol by capillary electrophoresis (CE). Chromate and the aromatics were separated in a relativ...

  2. Interfacing capillary electrophoresis with inductively coupled plasma mass spectrometry by direct injection nebulization for selenium speciation

    DEFF Research Database (Denmark)

    Bendahl, Lars; Gammelgaard, Bente; Jons, O.

    2001-01-01

    A demountable direct injection high efficiency nebulizer operating at low sample uptake rates was developed and used for coupling of capillary electrophoresis (CE) with inductively coupled plasma mass spectrometry (ICP-MS). When the nebulizer was used for continuous sample introduction, detection...

  3. Improving the reproducibility in capillary electrophoresis by incorporating current drift in mobility and peak area calculations

    DEFF Research Database (Denmark)

    Petersen, Nickolaj J.; Hansen, Steen H

    2012-01-01

    The traditional way of calculating mobility and peak areas in capillary electrophoresis does not take into account the changes in the buffer viscosity at different thermostatic control and that the analytes may accelerate during the individual runs due to Joule heating effects. We present a method...

  4. System zones in capillary zone electrophoresis: Moving boundaries caused by freely migrating hydroxide ions

    Czech Academy of Sciences Publication Activity Database

    Beckers, J. L.; Urbánek, Marek; Boček, Petr

    2005-01-01

    Roč. 26, č. 10 (2005), s. 1869-1873 ISSN 0173-0835 R&D Projects: GA AV ČR IAA4031103 Institutional research plan: CEZ:AV0Z40310501 Keywords : background electrolyte * capillary electrophoresis * system zone s Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.850, year: 2005

  5. IDENTIFICATION OF DEGRADATION PRODUCTS OF SOME CHEMICAL WARFARE AGENTS BY CAPILLARY ELECTROPHORESIS IONSPRAY MASS-SPECTROMETRY

    NARCIS (Netherlands)

    KOSTIAINEN, R; BRUINS, AP; HAKKINEN, VMA

    1993-01-01

    Capillary zone electrophoresis-ionspray mass spectrometry (CZE-IS-MS) in the negative-ion mode was applied in the identification of five organophosphonic acids, which are the primary hydrolysis products of nerve agents. The spectra exhibit a very abundant (M - H)- ion with minimal fragmentation.

  6. Microchip capillary electrophoresis for point-of-care analysis of lithium

    NARCIS (Netherlands)

    Vrouwe, E.X.; Luttge, R.; Vermes, I.; Berg, van den A.

    2007-01-01

    Background: Microchip capillary electrophoresis (CE) is a promising method for chemical analysis of complex samples such as whole blood. We evaluated the method for point-of-care testing of lithium. Methods: Chemical separation was performed on standard glass microchip CE devices with a conductivity

  7. Capillary electrophoresis in an extended nanospray tip-electrospray as an electrophoretic column

    Czech Academy of Sciences Publication Activity Database

    Týčová, Anna; Foret, František

    2015-01-01

    Roč. 1388, APR (2015), s. 274-279 ISSN 0021-9673 R&D Projects: GA ČR(CZ) GBP206/12/G014 Institutional support: RVO:68081715 Keywords : mass spectrometry * interface * separation * capillary electrophoresis Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.926, year: 2015

  8. Estimation of acidity constants, ionic mobilities and charges of antimicrobial peptides by capillary electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Tůmová, Tereza; Monincová, Lenka; Čeřovský, Václav; Kašička, Václav

    2016-01-01

    Roč. 37, 23/24 (2016), s. 3186-3195 ISSN 0173-0835 R&D Projects: GA ČR(CZ) GA15-01948S Institutional support: RVO:61388963 Keywords : acid dissociation constant * antimicrobial peptides * capillary electrophoresis * charge * mobility * Pka Subject RIV: CB - Analytical Chemistry , Separation Impact factor: 2.744, year: 2016

  9. Development of bufferless gel electrophoresis chip for easy preparation and rapid DNA separation.

    Science.gov (United States)

    Oleksandrov, Sergiy; Aman, Abdurazak; Lim, Wanyoung; Kim, Younghee; Bae, Nam Ho; Lee, Kyoung G; Lee, Seok Jae; Park, Sungsu

    2018-02-01

    This work presents a handy, fast, and compact bufferless gel electrophoresis chip (BGEC), which consists of precast agarose gel confined in a disposable plastic body with electrodes. It does not require large volumes of buffer to fill reservoirs, or the process of immersing the gel in the buffer. It withstands voltages up to 28.4 V/cm, thereby allowing DNA separation within 10 min with a similar separation capability to the standard gel electrophoresis. The results suggest that our BGEC is highly suitable for in situ gel electrophoresis in forensic, epidemiological settings and crime scenes where standard gel electrophoresis equipment cannot be brought in while quick results are needed. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. Nonaqueous capillary electrophoresis of dextromethorphan and its metabolites.

    Science.gov (United States)

    Pelcová, Marta; Langmajerová, Monika; Cvingráfová, Eliška; Juřica, Jan; Glatz, Zdeněk

    2014-10-01

    This study deals with the nonaqueous capillary electrophoretic separation of dextromethorphan and its metabolites using a methanolic background electrolyte. The optimization of separation conditions was performed in terms of the resolution of dextromethorphan and dextrorphan and the effect of separation temperature, voltage, and the characteristics of the background electrolyte were studied. Complete separation of all analytes was achieved in 40 mM ammonium acetate dissolved in methanol. Hydrodynamic injection was performed at 3 kPa for 4 s. The separation voltage was 20 kV accompanied by a low electric current. The ultraviolet detection was performed at 214 nm, the temperature of the capillary was 25°C. These conditions enabled the separation of four analytes plus the internal standard within 9 min. Further, the developed method was validated in terms of linearity, sensitivity, and repeatability. Rat liver perfusate samples were subjected to the nonaqueous capillary electrophoretic method to illustrate its applicability. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. Automation and integration of multiplexed on-line sample preparation with capillary electrophoresis for DNA sequencing

    Energy Technology Data Exchange (ETDEWEB)

    Tan, H.

    1999-03-31

    The purpose of this research is to develop a multiplexed sample processing system in conjunction with multiplexed capillary electrophoresis for high-throughput DNA sequencing. The concept from DNA template to called bases was first demonstrated with a manually operated single capillary system. Later, an automated microfluidic system with 8 channels based on the same principle was successfully constructed. The instrument automatically processes 8 templates through reaction, purification, denaturation, pre-concentration, injection, separation and detection in a parallel fashion. A multiplexed freeze/thaw switching principle and a distribution network were implemented to manage flow direction and sample transportation. Dye-labeled terminator cycle-sequencing reactions are performed in an 8-capillary array in a hot air thermal cycler. Subsequently, the sequencing ladders are directly loaded into a corresponding size-exclusion chromatographic column operated at {approximately} 60 C for purification. On-line denaturation and stacking injection for capillary electrophoresis is simultaneously accomplished at a cross assembly set at {approximately} 70 C. Not only the separation capillary array but also the reaction capillary array and purification columns can be regenerated after every run. DNA sequencing data from this system allow base calling up to 460 bases with accuracy of 98%.

  12. Studies on irradiated lactate dehydrogenase using gel electrophoresis

    International Nuclear Information System (INIS)

    Huth, O.

    1981-01-01

    LDH in aqueous phosphate buffer solution was x-irradiated under nitrogen and dinitrogen oxide. The pre-treated samples were separated by SDS-phosphate gel electrophoresis. Colour yields of the protein bands were determined by planimetry following staining and photometric densitometry. It could be shown that aggregates formed up to the tetramer size were mainly covalently bonded and that negligent amounts of fragments are formed. No exact statements could be made concerning products formed after longer irradiation treatment. Measurements under UV-light also revealed an elevated base from the point of application to the band at the native LDH level; this supports the theory of a reduced migration path through intra-molecular bonding. No firm conclusions can be drawn from the process of staining because CBB itself leads to an increased background colouring between closely-lying protein bands. It could be shown that OH-radicals cause greater changes than H-radicals or solvated electrons since the native LDH peak diminished fastest when irradiated under dinitrogen oxide. With regard to radiation effect it could be demonstrated that a higher radiation dose was required for degradation of the LDH molecule than for its inactivation. These values approach each other with increasing concentration. This indicates that at low concentrations inactivation of the enzyme is mainly caused by conformational changes and at high concentrations by aggregation. Disulphide bridges were found to contribute to aggregate formation in 5-15% of the aggregates formed, this increasing linearly with concentration. (orig./MG) [de

  13. Detection of circular telomeric DNA without 2D gel electrophoresis.

    Science.gov (United States)

    Dlaska, Margit; Anderl, Conrad; Eisterer, Wolfgang; Bechter, Oliver E

    2008-09-01

    The end of linear chromosomes forms a lasso-like structure called the t-loop. Such t-loops resemble a DNA recombination intermediate, where the single-stranded 3' overhang is arrested in a stretch of duplex DNA. Presumably, such a t-loop can also be deleted via a recombination process. This would result in the occurrence of circular extrachromosomal telomeric DNA (t-circles), which are known to be abundantly present in immortal cells engaging the recombination-based alternative lengthening of telomeres pathway (ALT pathway). Little is known about the basic mechanism of telomeric recombination in these cells and what ultimately causes the generation of such t-circles. Current standard procedures for detecting these molecules involve 2D gel electrophoresis or electron microscopy. However, both methods are labor intense and sophisticated to perform. Here, we present a simpler, faster, and equally sensitive method for detecting t-circles. Our approach is a telomere restriction fragment assay that involves the enzymatic preservation of circular DNA with Klenow enzyme followed by Bal31 degradation of the remaining linear DNA molecules. We show that with this approach t-circles can be detected in ALT cell lines, whereas no t-circles are present in telomerase-positive cell lines. We consider our approach a valid method in which t-circle generation is the experimental readout.

  14. [Analysis of Cut-off Value in Screening of Thalassemia by Capillary Hemoglobin Electrophoresis for Pregnant Women from Shenzhen Region of China].

    Science.gov (United States)

    Huo, Mei; Wu, Wen-Yuan; Liu, Mei; Gan, Zhi-Biao; Mao, Wei-Yu; Lin, Rong-Yao; Liu, Ai-Qin; He, Gui-Rong

    2016-04-01

    To investigate the cut-off value in screening of thalassemia in pregnant women from Shenzhen region by capillary hemoglobin electrophoresis. The data of capillary hemoglobin electrophoresis and genetic diagnosis of thalassemia from 2122 examined prenatal women were retrospectively analyzed. Capillary hemoglobin electrophoresis and α-, β- genetic diagnosis of thalassemia were carried out for every woman. Hemoglobin electrophoresis was performed using Capillarys 2 full-automated electrophoresis instrument. Gap polymerase chain reaction and reverse dot blot were used for genetic diagnosis of thalassemia genotyping test. The cut-off value in screening of thalassemia was determined by receiver operating characteristic curve and next to analyze the value of HbA2 and HbF in screening of thalassemia using the decided cut-off value. The areas under the curve (AUC(Roc)) of HbA2 for diagnosis of α-, β- thalassemia were 0.75 and 0.981 respectively, and the AUC(Roc) of HbF for diagnosis of β-thalassemia was 0.787. When HbA2 ≤ 2.55 was taken as the cut-off value of HbA2 for diagnosis of α-thalassemia, the sensitivity, specificity, positive likelihood ratio (LR(+)) and negative likelihood ratio (LR(-)) were 89.5%, 54.8%, 1.98, 0.19 respectively. When HbA2 ≥3.9 was taken as the cut off value of HbA2 for diagnosis of β-thalassemia, the sensitivity, specificity, LR(+) and LR(-) were 96.1%, 99.8% 480.5, 0.04 respectively. When HbF ≥0.75 was taken as the cut off value of HbF for diagnosis of β-thalassemia, the sensitivity, specificity, LR(+) and LR(-) were 83.6%, 61.8% respectively. The cut-off value in screening of thalassemia by capillarys 2 full automated electrophoresis instrument is different from that of the traditional method of hemoglobin electrophoresis, such as cellulose acetate membrane electrophoresis and agarose gel electrophoresis. Each laboratory should establish their own respective cut off value.

  15. Separation of 2-aminobenzoic acid-derivatized glycosaminoglycans and asparagine-linked glycans by capillary electrophoresis.

    Science.gov (United States)

    Sato, Kae; Sato, Kiichi; Okubo, Akira; Yamazaki, Sunao

    2005-01-01

    A capillary electrophoresis method was developed for the analysis of oligosaccharides combined with derivatization with 2-aminobenzoic acid. Glycosaminoglycan delta-disaccharides were effectively resolved on a fused-silica capillary tube using 150 mM borate, pH 8.5, as a running electrolyte solution. This analytical method was applied to the identification of glycosaminoglycan in combination with enzymatic digestion. The separation of N-glycans or glucose-oligomers was performed with a phosphate buffer containing polyethylene glycol or borate as an electrolyte solution. This method is expected to be useful in the determination of oligosaccharide structures in a glycoprotein.

  16. Lodenafil carbonate tablets: optimization and validation of a capillary zone electrophoresis method

    OpenAIRE

    Codevilla, Cristiane F; Ferreira, Pâmela Cristina L; Sangoi, Maximiliano S; Fröehlich, Pedro Eduardo; Bergold, Ana Maria

    2012-01-01

    A simple capillary zone electrophoresis (CZE) method was developed and validated for the analysis of lodenafil carbonate in tablets. Response surface methodology was used for optimization of the pH and concentration of the buffer, applied voltage and temperature. The method employed 50 mmol L-1 borate buffer at pH 10 as background electrolyte with an applied voltage of 15 kV. The separation was carried out in a fused-silica capillary maintained at 32.5 ºC and the detection wavelength was 214 ...

  17. Peptide separation by capillary electrophoresis with ultraviolet detection: Some simple approaches to enhance detection sensitivity and resolution

    International Nuclear Information System (INIS)

    Surugau, Noumie L.

    2011-01-01

    Capillary electrophoresis (CE) is one of the leading separation technologies for analysis of water-soluble analytes. CE has many advantages over the more established methods such as liquid chromatography and gel electrophoresis particularly in rapid analysis, require very little sample, use less or no toxic organic solvent, high peak efficiency and ease of automation. Despite the many attractive advantages of CE, CE users continue to seek improvements particularly on detection sensitivity, resolution and selectivity. This paper presented several simple approaches to improve detection sensitivity using simple sample pre-concentration called field-enhanced sample injection (FESI) and chromatographic-based ZipTip C 18 pre-concentrator. Also, some improvements in the resolution of complex peptides mixture when using two strategies namely, capillary coating and manipulation of the hydrophobicity of peptides using perfluorinated acids as background electrolyte (BGE), which have anionic conjugate base forms with hydrophobic character. As test compounds, standard peptide mixture and proteins digests were used for these studies. The results showed that FESI has significantly enhanced the detection signal of peptide standards and bovine serum albumin (BSA) tryptic digests. As for the use of ZipTip C 18 pre-concentrator, selective enhancement in detection signal was particularly notable on the late migrating peptides. Coating the capillary proved to have little changes on the CE of peptides when used in conjunction with acidic BGE. Electropherograms of BSA tryptic peptides in pentafluoropropionic acid (PFPA) and heptafluorobutyric acid (HFBA) showed interesting profile, with notable resolution improvement for peptides with close similarity in electrophoretic mobilities. (author)

  18. Amplified fragment length polymorphism and pulsed field gel electrophoresis for subspecies differentiation of Serpulina pilosicoli

    DEFF Research Database (Denmark)

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

    1999-01-01

    Pulsed field gel electrophoresis (PFGE) and amplified fragment length polymorphism (AFLP) were compared for their ability to differentiate between 50 porcine Serpulina pilosicoli isolates. Both techniques were highly sensitive, dividing the isolates into 36 and 38 groups, respectively. Due...

  19. Ultrafast Capillary Electrophoresis Isolation of DNA Aptamer for the PCR Amplification-Based Small Analyte Sensing

    Directory of Open Access Journals (Sweden)

    Emmanuelle eFiore

    2015-08-01

    Full Text Available Here, we report a new homogeneous DNA amplification-based aptamer assay for small analyte sensing. The aptamer of adenosine chosen as the model analyte was split into two fragments able to assemble in the presence of target. Primers were introduced at extremities of one fragment in order to generate the amplifiable DNA component. The amount of amplifiable fragment was quantifiable by Real-Time Polymerase Chain Reaction (RT-PCR amplification and directly reliable on adenosine concentration. This approach combines the very high separation efficiency and the homogeneous format (without immobilization of capillary electrophoresis and the sensitivity of real time PCR amplification. An ultrafast isolation of target-bound split aptamer (60 s was developed by designing a capillary electrophoresis input/ouput scheme. Such method was successfully applied to the determination of adenosine with a LOD of 1 µM.

  20. Development of a multiplexed interface for capillary electrophoresis-electrospray ion trap mass spectrometry.

    Science.gov (United States)

    Li, Fu-An; Wu, Ming-Chi; Her, Guor-Rong

    2006-08-01

    A four-channel multiplexed electrospray capillary electrophoresis interface has been developed. This new interface permits up to four capillary electrophoresis columns to be sampled sequentially by means of a stepper motor and a notched rotating plate assembly, which at any instant occludes all but a single sprayer. In this design, four sheath liquid electrospray probes are oriented in a circular array situated 90 degrees relative to one another. The rotating metal disk, which contains a one-quarter notch, is mounted to the stepper motor assembly and is located between the sprayers and the entrance aperture of an ion trap mass spectrometer. By using the data acquisition signal from the ion trap mass spectrometer, the scan event is synchronized with the rotation of the metal disk. With this device, four discrete sample streams can be simultaneously analyzed, resulting in a 4-fold increase in analytical throughput.

  1. Simultaneous determination of five flavonoids in saussurea involucrata by capillary electrophoresis

    International Nuclear Information System (INIS)

    Li, Y.; Zhong, H.; Zhong, H.

    2013-01-01

    A method of determination of five flavonoids in Saussurea involucrata by beta-cyclodextrin modified capillary zone electrophoresis has been developed.The effects of buffer pH and buffer concentration, applied voltage and beta-CD concentrations on the separation were systematically investigated. The optimum condition providing baseline separation of all compounds within 8 min was obtained in the 20 mmol per liter borax buffer (pH 9.2), 20 kV applied voltage and 8 mmol per liter beta-CD. The linearity, detection limits, limits of quantification, reproducibility and recovery were satisfactory. The beta-cyclodextrin modified capillary zone electrophoresis method proposed here has been satisfactorily employed to analyze S. involucrate samples. (author)

  2. Determination of Na and Al Ions in Semiconductor Cleaning Solution Using Capillary Electrophoresis

    International Nuclear Information System (INIS)

    Lee, H. P.; Lim, H. B.

    2003-01-01

    The most common process chemical used in the manufacturing process is a standard cleaning (SC) solution, a mixture of ammonia and hydrogen peroxide in deionized water. Since the purity of the SC solution used in the process has been required to the level of sub-ppb range, accurate and reliable determination of ionic contaminants becomes increasingly difficult. In order to satisfy the requirement of impurity control, inductively coupled plasma-mass spectrometer (ICP-MS), graphite furnace atomic absorption spectrometer (GFAAS), and ion chromatography (IC) are currently the most common analytical instruments used in the process. However, those instruments are not designed for on-line monitoring but rather for off-line analysis. Recently, separation and detection of various particles, such as cells and nanoparticles, with capillary electrophoresis (CE) was reported, although the application of CE has been mostly limited to organic or biological samples. Capillary electrophoresis has been emerging as an alternative to ICPAES and AAS for trace metal analysis

  3. Analysis of arecoline in Semen Arecae decoction pieces by microchip capillary electrophoresis with contactless conductivity detection

    Directory of Open Access Journals (Sweden)

    Zi-You Cai

    2012-10-01

    Full Text Available A new method for the determination of arecoline in Semen Arecae decoction pieces by microchip capillary electrophoresis with contactless conductivity detection (MCE-CCD was proposed. The effects of various electrophoretic operating parameters on the analysis of arecoline were studied. Under the optimal conditions, arecoline was rapidly separated and detected in 1 min with good linearity over the concentration range of 20–1500 μM (r2=0.9991 and the detection limit of 5 μM (S/N=3. The method was used for the analysis of arecoline satisfactorily with a recovery of 96.8–104%. Keywords: Microchip capillary electrophoresis, Contactless conductivity detection, Arecoline, Semen Arecae

  4. The application of single cell gel electrophoresis or comet assay to human monitoring studies

    Directory of Open Access Journals (Sweden)

    Valverde Mahara

    1999-01-01

    Full Text Available Objective. In the search of new human genotoxic biomarkers, the single cell gel electrophoresis assay has been proposed as a sensible alternative. Material and methods. This technique detects principally single strand breaks as well as alkali-labile and repair-retarded sites. Results. Herein we present our experience using the single cell gel electrophoresis assay in human population studies, both occupationally and environmentally exposed. Conclusions. We discuss the assay feasibility as a genotoxic biomarker.

  5. Detection of human DNA polymorphisms with a simplified denaturing gradient gel electrophoresis technique.

    OpenAIRE

    Noll, W W; Collins, M

    1987-01-01

    Single base pair differences between otherwise identical DNA molecules can result in altered melting behavior detectable by denaturing gradient gel electrophoresis. We have developed a simplified procedure for using denaturing gradient gel electrophoresis to detect base pair changes in genomic DNA. Genomic DNA is digested with restriction enzymes and hybridized in solution to labeled single-stranded probe DNA. The excess probe is then hybridized to complementary phage M13 template DNA, and th...

  6. Electromembrane extraction of heavy metal cations followed by capillary electrophoresis with capacitively coupled contactless conductivity detection

    Czech Academy of Sciences Publication Activity Database

    Kubáň, Pavel; Strieglerová, Lenka; Gebauer, Petr; Boček, Petr

    2011-01-01

    Roč. 32, č. 9 (2011), s. 1025-1032 ISSN 0173-0835 R&D Projects: GA ČR GA203/08/1536; GA ČR GAP206/10/1219; GA AV ČR IAA400310703 Institutional research plan: CEZ:AV0Z40310501 Keywords : capillary electrophoresis * electromembrane extraction * heavy metal cations Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.303, year: 2011

  7. Photodeposited silver nanoparticles for on-column surface-enhanced Raman spectrometry detection in capillary electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Přikryl, Jan; Klepárník, Karel; Foret, František

    2012-01-01

    Roč. 1226, - (2012), s. 43-47 ISSN 0021-9673 R&D Projects: GA ČR GA203/08/1680; GA ČR(CZ) GAP301/11/2055; GA MŠk LC06023 Institutional research plan: CEZ:AV0Z40310501 Keywords : SERS * capillary electrophoresis * photodeposition Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 4.612, year: 2012

  8. Determination of gamma-hydroxybutyric acid in clinical samples using capillary electrophoresis with contactless conductivity detection

    Czech Academy of Sciences Publication Activity Database

    Gong, X.Y.; Kubáň, Pavel; Scholer, A.; Hauser, P.C.

    2008-01-01

    Roč. 1213, 1-2 (2008), s. 100-104 ISSN 0021-9673 R&D Projects: GA AV ČR IAA400310609; GA AV ČR IAA400310703; GA ČR GA203/08/1536 Institutional research plan: CEZ:AV0Z40310501 Keywords : clinical samples * capillary electrophoresis * contactless conductivity detection Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.756, year: 2008

  9. Determination of uric acid in plasma and allantoic fluid of chicken embryos by capillary electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Matějčková, J.; Tůma, P.; Samcová, E.; Zemanová, Zdeňka

    2007-01-01

    Roč. 30, č. 12 (2007), s. 1947-1952 ISSN 1615-9306 R&D Projects: GA ČR(CZ) GA304/04/0972 Grant - others:GA ČR(CZ) GA203/07/0896 Program:GA Institutional research plan: CEZ:AV0Z50110509 Keywords : capillary electrophoresis * microsamples of plasma * uric acid Subject RIV: CE - Biochemistry Impact factor: 2.632, year: 2007

  10. Determination of dissociation constants of compounds with potential cognition enhancing activity by capillary zone electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Lišková, A.; Křivánková, Ludmila

    2005-01-01

    Roč. 26, č. 23 (2005), s. 4429-4439 ISSN 0173-0835 R&D Projects: GA ČR GA203/05/2106; GA AV ČR IAA4031401 Institutional research plan: CEZ:AV0Z40310501 Keywords : Capillary zone electrophoresis * dissociation constants * potential nootropics Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.850, year: 2005

  11. Determination of acid dissociation constants of triazole fungicides by pressure assisted capillary electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Konášová, Renáta; Jaklová Dytrtová, Jana; Kašička, Václav

    2015-01-01

    Roč. 1408, Aug 21 (2015), s. 243-249 ISSN 0021-9673 R&D Projects: GA ČR(CZ) GA13-17224S; GA ČR GP13-21409P Institutional support: RVO:61388963 Keywords : triazole fungicides * acid dissociation constant * pK(a) * capillary electrophoresis * ionic mobility Subject RIV: CB - Analytical Chemistry , Separation Impact factor: 3.926, year: 2015

  12. Capillary electrophoresis as a screening tool for alpha amylase inhibitors in plant extracts

    OpenAIRE

    Hamdan, Imad I.; Afifi, Fatima U.

    2010-01-01

    Capillary electrophoresis (CE) method was developed for screening plant extract for potential alpha amylase (AA) inhibitory activity. The method was validated against a well established UV method. Overall, the proposed method was shown able to detect plants with significant alpha amylase inhibitory activity but not those with rather clinically insignificant activities. Fifty plant species were screened using both the proposed CE method and the UV method and seven plant species were found to p...

  13. Enantioselective analysis of drugs: contributions of high-performance liquid chromatography and capillary electrophoresis

    OpenAIRE

    Bonato, Pierina Sueli; Jabor, Valquíria Aparecida Polisel; Gaitani, Cristiane Masetto de

    2005-01-01

    The demand for analytical methods suitable for accurate and reproducible determination of drug enantiomers has increased significantly in the last years. High-performance liquid chromatography (HPLC) using chiral stationary phases and capillary electrophoresis (CE) are the most important techniques used for this purpose. In this paper, the fundamental aspects of chiral separations using both techniques are presented. Some important aspects for the development of enantioselective methods, part...

  14. Why robust background electrolytes containing multivalent ionic species can fail in capillary zone electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Beckers, J. L.; Gebauer, Petr; Boček, Petr

    2001-01-01

    Roč. 916, č. 1 (2001), s. 41-49 ISSN 0021-9673 R&D Projects: GA AV ČR IAA4031703; GA AV ČR IAA4031103; GA ČR GA203/99/0044; GA ČR GA203/01/0401 Institutional research plan: CEZ:AV0Z4031919 Keywords : capillary zone electrophoresis * system peak s * organic acids Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 2.793, year: 2001

  15. Determination of monosaccharides derivatized with 2-aminobenzoic Acid by capillary electrophoresis.

    Science.gov (United States)

    Abo, Mitsuru; He, Li-Ping; Sato, Kae; Okubo, Akira

    2013-01-01

    Reducing monosaccharides were derivatized with 2-aminobenzoic acid (2-AA) through reductive amination using sodium cyanoborohydride as a reductant, and the derivatives were separated by capillary zone electrophoresis with UV detection using 50 mM sodium phosphate (pH 5.5) or 150 mM sodium borate-50 mM sodium phosphate (pH 7.0) running buffer. The derivatives of monosaccharides, which are major components of various carbohydrate materials, were completely separated within 25 min.

  16. Evaluation of carrier ampholyte-based capillary electrophoresis for separation of peptides and peptide mimetics

    Czech Academy of Sciences Publication Activity Database

    Koval, Dušan; Busnel, J. M.; Hlaváček, Jan; Jiráček, Jiří; Kašička, Václav; Peltre, G.

    2008-01-01

    Roč. 29, č. 18 (2008), s. 3759-3767 ISSN 0173-0835 R&D Projects: GA ČR(CZ) GA203/06/1044; GA ČR GA203/06/1272; GA ČR GA203/08/1428 Institutional research plan: CEZ:AV0Z40550506 Keywords : capillary electrophoresis * carrier ampholytes * peptides Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.509, year: 2008

  17. System zones in capillary zone electrophoresis: Moving boundaries caused by freely migrating hydrogen ions

    Czech Academy of Sciences Publication Activity Database

    Beckers, J. L.; Boček, Petr

    2005-01-01

    Roč. 26, č. 2 (2005), s. 446-452 ISSN 0173-0835 R&D Projects: GA ČR(CZ) GA203/02/0023; GA AV ČR(CZ) IAA4031401; GA AV ČR(CZ) IAA4031103 Institutional research plan: CEZ:AV0Z40310501 Keywords : capillary zone electrophoresis * system zone s Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.850, year: 2005

  18. Interactions of helquats with chiral acidic aromatic analytes investigated by partial-filling affinity capillary electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Růžička, Martin; Koval, Dušan; Vávra, Jan; Reyes Gutierrez, Paul Eduardo; Teplý, Filip; Kašička, Václav

    2016-01-01

    Roč. 1467, Oct 7 (2016), s. 417-426 ISSN 0021-9673 R&D Projects: GA ČR(CZ) GA15-01948S; GA ČR GA13-32974S; GA ČR GA13-19213S Institutional support: RVO:61388963 Keywords : affinity capillary electrophoresis * binding constant * chiral separation * helquats * noncovalent interactions * partial filling Subject RIV: CB - Analytical Chemistry , Separation Impact factor: 3.981, year: 2016

  19. Steroid determination in fish plasma using capillary electrophoresis

    Science.gov (United States)

    Bykova, L.; Archer-Hartmann, S. A.; Holland, L.A.; Iwanowicz, L.R.; Blazer, V.S.

    2010-01-01

    A capillary separation method that incorporates pH-mediated stacking is employed for the simultaneous determination of circulating steroid hormones in plasma from Perca flavescens (yellow perch) collected from natural aquatic environments. The method can be applied to separate eight steroid standards: progesterone, 17α,20β-dihydroxypregn-4-en-3-one, 17α-hydroxyprogesterone, testosterone, estrone, 11-ketotestosterone, ethynyl estradiol, and 17β-estradiol. Based on screening of plasma, the performance of the analytical method was determined for 17α,20β-dihydroxypregn-4-en-3-one, testosterone, 11-ketotestosterone, and 17β-estradiol. The within-day reproducibility in migration time for these four steroids in aqueous samples was ≤2%. Steroid quantification was accomplished using a calibration curve obtained with external standards. Plasma samples from fish collected from the Choptank and Severn Rivers, Maryland, USA, stored for up to one year were extracted with ethyl acetate and then further processed with anion exchange and hydrophobic solid phase extraction cartridges. The recovery of testosterone and 17β-estradiol from yellow perch plasma was 84 and 85%, respectively. Endogenous levels of testosterone ranged from 0.9 to 44 ng/ml, and when detected 17α,20β-dihydroxypregn-4-en-3-one ranged from 5 to 34 ng/ml. The reported values for testosterone correlated well with the immunoassay technique. Endogenous concentrations of 17β-estradiol were ≤1.7 ng/ml. 11-Ketotestosterone was not quantified because of a suspected interferant. Higher levels of 17α,20β-dihydroxypregn-4-en-3-one were found in male and female fish in which 17β-estradiol was not detected. Monitoring multiple steroids can provide insight into hormonal fluctuations in fish.

  20. Steroid determination in fish plasma using capillary electrophoresis.

    Science.gov (United States)

    Bykova, Liliya; Archer-Hartmann, Stephanie A; Holland, Lisa A; Iwanowicz, Luke R; Blazer, Vicki S

    2010-09-01

    A capillary separation method that incorporates pH-mediated stacking is employed for the simultaneous determination of circulating steroid hormones in plasma from Perca flavescens (yellow perch) collected from natural aquatic environments. The method can be applied to separate eight steroid standards: progesterone, 17alpha,20beta-dihydroxypregn-4-en-3-one, 17alpha-hydroxyprogesterone, testosterone, estrone, 11-ketotestosterone, ethynyl estradiol, and 17beta-estradiol. Based on screening of plasma, the performance of the analytical method was determined for 17alpha,20beta-dihydroxypregn-4-en-3-one, testosterone, 11-ketotestosterone, and 17beta-estradiol. The within-day reproducibility in migration time for these four steroids in aqueous samples was < or =2%. Steroid quantification was accomplished using a calibration curve obtained with external standards. Plasma samples from fish collected from the Choptank and Severn Rivers, Maryland, USA, stored for up to one year were extracted with ethyl acetate and then further processed with anion exchange and hydrophobic solid phase extraction cartridges. The recovery of testosterone and 17beta-estradiol from yellow perch plasma was 84 and 85%, respectively. Endogenous levels of testosterone ranged from 0.9 to 44 ng/ml, and when detected 17alpha,20beta-dihydroxypregn-4-en-3-one ranged from 5 to 34 ng/ml. The reported values for testosterone correlated well with the immunoassay technique. Endogenous concentrations of 17beta-estradiol were < or =1.7 ng/ml. 11-Ketotestosterone was not quantified because of a suspected interferant. Higher levels of 17alpha,20beta-dihydroxypregn-4-en-3-one were found in male and female fish in which 17beta-estradiol was not detected. Monitoring multiple steroids can provide insight into hormonal fluctuations in fish. Copyright 2010 SETAC.

  1. A Novel Protocol to Analyze Short- and Long-Chain Fatty Acids Using Nonaqueous Microchip Capillary Electrophoresis

    Science.gov (United States)

    Cable, M. L.; Stockton, A. M.; Mora, Maria F; Willis, P. A.

    2013-01-01

    We propose a new protocol to identify and quantify both short- and long-chain saturated fatty acids in samples of astrobiological interest using non-aqueous microchip capillary electrophoresis (micronNACE) with laser induced fluorescence (LIF).

  2. Determination of acid dissociation constants of warfarin and hydroxywarfarins by capillary electrophoresis.

    Science.gov (United States)

    Nowak, Paweł; Olechowska, Paulina; Mitoraj, Mariusz; Woźniakiewicz, Michał; Kościelniak, Paweł

    2015-08-10

    In this work the acid dissociation constants--pKa of warfarin and its all important oxidative metabolites have been determined by capillary electrophoresis-based methods. It has resulted in a complete description of two acid-base dissociation equilibria, yet not investigated experimentally for phase I metabolites of warfarin. The capillary electrophoresis (CE) method based on the relation between effective electrophoretic mobilities and pH has proven to be a suitable tool for pKa determination, while the spectrophotometric (CE-DAD) and the internal standard methods (IS-CE), have appeared to be promising alternative approaches. The CE-DAD approach based on the change in absorbance spectra between the acidic and basic forms is a combination between capillary electrophoresis and spectrophotometric titration, and yields very consistent values of pKa1 with CE. The IS-CE, in turn, enables an estimation of pKa1 and pKa2 from only two analytical runs, however, less accurate than CE and CE-DAD. The Debye-Hückel model has been confirmed experimentally as a good predictor of pKa values at various ionic strengths. Therefore, it has been used in determination of thermodynamic pKa1 and pKa2, referring to the zero ionic strength. The results are important from the analytical, pharmacological, and theoretical points of view. Copyright © 2015 Elsevier B.V. All rights reserved.

  3. Sensitive detection of malachite green and crystal violet by nonlinear laser wave mixing and capillary electrophoresis.

    Science.gov (United States)

    Maxwell, Eric J; Tong, William G

    2016-05-01

    An ultrasensitive label-free antibody-free detection method for malachite green and crystal violet is presented using nonlinear laser wave-mixing spectroscopy and capillary zone electrophoresis. Wave-mixing spectroscopy provides a sensitive absorption-based detection method for trace analytes. This is accomplished by forming dynamic gratings within a sample cell, which diffracts light to create a coherent laser-like signal beam with high optical efficiency and high signal-to-noise ratio. A cubic dependence on laser power and square dependence on analyte concentration make wave mixing sensitive enough to detect molecules in their native form without the use of fluorescent labels for signal enhancement. A 532 nm laser and a 635 nm laser were used for malachite green and crystal violet sample excitation. The use of two lasers of different wavelengths allows the method to simultaneously detect both analytes. Selectivity is obtained through the capillary zone electrophoresis separation, which results in characteristic migration times. Measurement in capillary zone electrophoresis resulted in a limit of detection of 6.9 × 10(-10)M (2.5 × 10(-19) mol) for crystal violet and 8.3 × 10(-11)M (3.0 × 10(-20) mol) for malachite green at S/N of 2. Copyright © 2016. Published by Elsevier B.V.

  4. Detection system of capillary array electrophoresis microchip based on optical fiber

    Science.gov (United States)

    Yang, Xiaobo; Bai, Haiming; Yan, Weiping

    2009-11-01

    To meet the demands of the post-genomic era study and the large parallel detections of epidemic diseases and drug screening, the high throughput micro-fluidic detection system is needed urgently. A scanning laser induced fluorescence detection system based on optical fiber has been established by using a green laser diode double-pumped solid-state laser as excitation source. It includes laser induced fluorescence detection subsystem, capillary array electrophoresis micro-chip, channel identification unit and fluorescent signal processing subsystem. V-shaped detecting probe composed with two optical fibers for transmitting the excitation light and detecting induced fluorescence were constructed. Parallel four-channel signal analysis of capillary electrophoresis was performed on this system by using Rhodamine B as the sample. The distinction of different samples and separation of samples were achieved with the constructed detection system. The lowest detected concentration is 1×10-5 mol/L for Rhodamine B. The results show that the detection system possesses some advantages, such as compact structure, better stability and higher sensitivity, which are beneficial to the development of microminiaturization and integration of capillary array electrophoresis chip.

  5. A two-step method for rapid characterization of electroosmotic flows in capillary electrophoresis.

    Science.gov (United States)

    Zhang, Wenjing; He, Muyi; Yuan, Tao; Xu, Wei

    2017-12-01

    The measurement of electroosmotic flow (EOF) is important in a capillary electrophoresis (CE) experiment in terms of performance optimization and stability improvement. Although several methods exist, there are demanding needs to accurately characterize ultra-low electroosmotic flow rates (EOF rates), such as in coated capillaries used in protein separations. In this work, a new method, called the two-step method, was developed to accurately and rapidly measure EOF rates in a capillary, especially for measuring the ultra-low EOF rates in coated capillaries. In this two-step method, the EOF rates were calculated by measuring the migration time difference of a neutral marker in two consecutive experiments, in which a pressure driven was introduced to accelerate the migration and the DC voltage was reversed to switch the EOF direction. Uncoated capillaries were first characterized by both this two-step method and a conventional method to confirm the validity of this new method. Then this new method was applied in the study of coated capillaries. Results show that this new method is not only fast in speed, but also better in accuracy. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. Thalassemia and hemoglobinopathies in Southeast Asian newborns: diagnostic assessment using capillary electrophoresis system.

    Science.gov (United States)

    Srivorakun, Hataichanok; Fucharoen, Goonnapa; Changtrakul, Yossombat; Komwilaisak, Patcharee; Fucharoen, Supan

    2011-04-01

    We have investigated the Capillarys 2 Hemoglobin testing system to assist in presumptive diagnosis of thalassemia and hemoglobinopathies commonly found in Southeast Asia. Study was conducted on 226 newborns. Hematological parameters were recorded and Hb profiles were examined on the Capillarys 2 Hemoglobin analyzer (SEBIA). DNA analyses were used to establish the final diagnoses. Among 226 newborns examined, 122 had thalassemias with 17 different genotypes. The capillary electrophoresis system could provide useful data for presumptive diagnoses of cases, especially those with Hb E and α-thalassemia. Hb E was found to be 2.6-6.2% in heterozygote whereas Hb Bart's were clearly observed in cases with compound heterozygous or homozygous α(+)-thalassemia and heterozygous α(0)-thalassemia. Hb H disease and other forms of α-thalassemia could be differentiated based on the presence of Hb Bart's and its percentage. The capillary electrophoresis system is applicable to newborn screening for common forms of thalassemia in Southeast Asia. Copyright © 2011 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

  7. ssDNA degradation along capillary electrophoresis process using a Tris buffer.

    Science.gov (United States)

    Ric, Audrey; Ong-Meang, Varravaddheay; Poinsot, Verena; Martins-Froment, Nathalie; Chauvet, Fabien; Boutonnet, Audrey; Ginot, Frédéric; Ecochard, Vincent; Paquereau, Laurent; Couderc, François

    2017-06-01

    Tris-Acetate buffer is currently used in the selection and the characterization of ssDNA by capillary electrophoresis (CE). By applying high voltage, the migration of ionic species into the capillary generates a current that induces water electrolysis. This phenomenon is followed by the modification of the pH and the production of Tris derivatives. By injecting ten times by capillary electrophoresis ssDNA (50 nM), the whole oligonucleotide was degraded. In this paper, we will show that the Tris buffer in the running vials is modified along the electrophoretic process by electrochemical reactions. We also observed that the composition of the metal ions changes in the running buffer vials. This phenomenon, never described in CE, is important for fluorescent ssDNA analysis using Tris buffer. The oligonucleotides are degraded by electrochemically synthesized species (present in the running Tris vials) until it disappears, even if the separation buffer in the capillary is clean. To address these issues, we propose to use a sodium phosphate buffer that we demonstrate to be electrochemically inactive. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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

    DEFF Research Database (Denmark)

    Doll, Hans; Andersen, Bente

    1981-01-01

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

  9. Capillary electrophoresis fragment analysis and clone sequencing in detection of dynamic mutations of spinocerebellar ataxia

    Directory of Open Access Journals (Sweden)

    Yuan-yuan CHEN

    2018-04-01

    Full Text Available Objective To estimate the accuracy and stability of capillary electrophoresis fragment analysis and clone sequencing in detecting dynamic mutations of spinocerebellar ataxia (SCA. Methods Capillary electrophoresis fragment analysis and clone sequencing were used in detecting trinucleotide repeated sequence of 14 SCA patients (3 cases of SCA2, 2 cases of SCA7, 7 cases of SCA8 and 2 cases of SCA17. Results Capillary electrophoresis fragment analysis of 3 SCA2 cases showed the expanded cytosine-adenine-guanine (CAG repeats were 31, 30 and 32, and the copy numbers of 3 clone sequencing for 3 colonies in each case were 37/40/40, 37/38/39 and 38/39/40 respectively. Capillary electrophoresis fragment analysis of 2 SCA7 cases showed the expanded CAG repeats were 57 and 34, and the copy numbers of repeats were 69, 74, 75 in 3 colonies of one case, and was 45 in the other case. For the 7 SCA8 cases with the expanded cytosine-thymine-adenine (CTA/cytosine-thymine-guanine (CTG repeats of 99, 111, 104, 92, 89, 104 and 75, the results of clone sequencing were 97, 116, 104, 90, 90, 102 and 76 respectively. For 2 SCA17 cases with the short/expanded CAG repeats of 37/50 and 36/45, the results of clone sequencing were 51/50/52 and 45/44 for 3 and 2 colonies. Conclusions Although the higher mobility of polymerase chain reaction (PCR products containing dynamic mutation in the capillary electrophoresis fragment analysis might cause the deviation for analysis of copy numbers, the deviation was predictable and the results were repeatable. The clone sequencing results showed obvious instability, especially for SCA2 and SCA7 genes, which might owing to their simple CAG repeats. Consequently, clone sequencing is not suited for detection of dynamic mutation, not to mention the quantitative criteria of dynamic mutation sequencing. DOI: 10.3969/j.issn.1672-6731.2018.03.008

  10. A Theoretical Analysis of the Influence of Electroosmosis on the Effective Ionic Mobility in Capillary Zone Electrophoresis

    Science.gov (United States)

    Hijnen, Hens

    2009-01-01

    A theoretical description of the influence of electroosmosis on the effective mobility of simple ions in capillary zone electrophoresis is presented. The mathematical equations derived from the space-charge model contain the pK[subscript a] value and the density of the weak acid surface groups as parameters characterizing the capillary. It is…

  11. The characteristics of transferrin variants by carbohydrate-deficient transferrin tests using capillary zone electrophoresis.

    Science.gov (United States)

    Yoo, Gilsung; Kim, Juwon; Yoon, Kap Joon; Lee, Jong-Han

    2018-04-17

    Transferrin is the major plasma transport protein for iron. We aimed to investigate the characteristics of transferrin variant by carbohydrate-deficient transferrin (CDT) test using capillary zone electrophoresis. We retrospectively analyzed the CDT tests of 2449 patients from March 2009 to May 2017 at a tertiary hospital in Korea. CDT was quantified using a Capillarys 2 system (Sebia, Lisses, France) by capillary zone electrophoresis. The characteristics of variant transferrin patterns using electropherogram of CDT tests were analyzed. Seventy-seven (3.1%) patients were classified as variant transferrin. Mean age of these patients was 51.8 years, and the male-to-female ratio was 3.5:1. The most common variants were the BC variants (n = 37), followed by the CD variants (n = 27), unclear patterns (n = 7), BD variants (n = 3), CC variants (n = 2), misclassification (n = 1). In the variant Tf group, the most common disease was alcoholic liver cirrhosis (n = 22, 28.6%), followed by the toxic effects of substances (n = 17, 22.1%), and mental and behavioral disorders attributable to alcohol (n = 11, 14.3%). Nonvariant group showed a predominance of the toxic substance effects (n = 880, 37.1%), a personal history of suicide attempts (n = 634, 26.7%), and mental and behavioral disorders due to alcohol (n = 336, 14.2%). We analyzed the basic characteristics of variant transferrin by CDT tests using capillary zone electrophoresis. The prevalence of variant transferrin was 3.1% of the study subjects. Male patients, alcohol abusers, and liver cirrhosis patients predominated in the variant transferrin population. Further prospective studies are warranted to elucidate variant transferrin in clinical practice. © 2018 Wiley Periodicals, Inc.

  12. Developments in coupled solid-phase extraction-capillary electrophoresis 2013-2015.

    Science.gov (United States)

    Ramautar, Rawi; Somsen, Govert W; de Jong, Gerhardus J

    2016-01-01

    An overview of the design and application of coupled solid-phase extraction-capillary electrophoresis (SPE-CE) systems reported in the literature between July 2013 and June 2015 is provided in this paper. The present article is a continuation of our previous review papers on this topic which covered the time period 2000-2013 (Electrophoresis 2008, 29, 108-128; Electrophoresis 2010, 31, 44-54; Electrophoresis 2012, 33, 243-250; Electrophoresis 2014, 35, 128-137). The use of in-line and on-line SPE-CE approaches is treated and outlined in this review. Recent advancements, such as, for example, the use of aptamers as affinity material for in-line SPE-CE, the use of a bead string design for in-line fritless SPE-CE, and new interfacing techniques for the on-line coupling of SPE to CE, are outlined. Selected examples demonstrate the applicability of the coupled SPE-CE systems for biomedical, pharmaceutical, environmental, and food studies. A complete overview of the recent SPE-CE studies is given in table format, providing information on sample type, SPE sorbent, coupling mode, detection mode, and LOD. Finally, some general conclusions and perspectives are provided. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Adapting mass spectrometry-based platforms for clinical proteomics applications: The capillary electrophoresis coupled mass spectrometry paradigm

    Science.gov (United States)

    Metzger, Jochen; Luppa, Peter B.; Good, David M.; Mischak, Harald

    2018-01-01

    Single biomarker detection is common in clinical laboratories due to the currently available method spectrum. For various diseases, however, no specific single biomarker could be identified. A strategy to overcome this diagnostic void is to shift from single analyte detection to multiplexed biomarker profiling. Mass spectrometric methods were employed for biomarker discovery in body fluids. The enormous complexity of biofluidic proteome compartments implies upstream fractionation. For this reason, mass spectrometry (MS) was coupled to two-dimensional gel electrophoresis, liquid chromatography, surface-enhanced laser desorption/ionization, or capillary electrophoresis (CE). Differences in performance and operating characteristics make them differentially suited for routine laboratory applications. Progress in the field of clinical proteomics relies not only on the use of an adequate technological platform, but also on a fast and efficient proteomic workflow including standardized sample preparation, proteomic data processing, statistical validation of biomarker selection, and sample classification. Based on CE-MS analysis, we describe how proteomic technology can be implemented in a clinical laboratory environment. In the last part of this review, we give an overview of CE-MS-based clinical studies and present information on identity and biological significance of the identified peptide biomarkers providing evidence of disease-induced changes in proteolytic processing and posttranslational modification. PMID:19404829

  14. Sheath-flow electrochemical detection of amino acids with a copper wire electrode in capillary electrophoresis.

    Science.gov (United States)

    Inoue, Junji; Kaneta, Takashi; Imasaka, Totaro

    2012-09-01

    Here, we report the detection of native amino acids using a sheath-flow electrochemical detector with a working electrode made of copper wire. A separation capillary that was inserted into a platinum tube in the detector acted as a grounded electrode for electrophoresis and as a flow channel for sheath liquid. Sheath liquid flowed outside the capillary to support the transport of the separated analytes to the working electrode for electrochemical detection. The copper wire electrode was aligned at the outlet of the capillary in a wall-jet configuration. Amino acids injected into the capillary were separated following elution from the end of the capillary and detection by the copper electrode. Three kinds of copper electrodes with different diameters-50, 125, and 300 μm-were examined to investigate the effect of the electrode diameter on sensitivity. The peak widths of the analytes were independent of the diameter of the working electrode, while the 300-μm electrode led to a decrease in the signal-to-noise ratio compared with the 50- and 125-μm electrodes, which showed no significant difference. The flow rate of the sheath liquid was also varied to optimize the detection conditions. The limits of detection for amino acids ranged from 4.4 to 27 μM under optimal conditions. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Application of capillary electrophoresis to anion speciation in soil water extracts: 2. Arsenic

    Energy Technology Data Exchange (ETDEWEB)

    Naidu, R.; Smith, J.; McLaren, R.G.; Stevens, D.P.; Sumner, M.E.; Jackson, P.E.

    2000-02-01

    A method has been developed for the speciation of arsenic (AsO{sub 2}{sup {minus}}, AsO{sub 4}{sup 3{minus}}, and dimethylarsinic [DMA]) in natural soil solutions from contaminated sites in Australia. The separation of these anions was achieved by capillary zone electrophoresis (CZE) using a fused silica capillary with a basic chromate buffer and on-column indirect UV detection at 254 nm. Method parameters, such as electrolyte pH, run voltage, and capillary temperature were studied in order to establish suitable analytical conditions. The ideal separation for As(III) and DMA was achieved with a buffer pH of 8.0, a run voltage of 25 kV, and a capillary temperature of 30 C. Under these conditions, As(V) and orthophosphate ions comigrated. However, the use of a chromate buffer at pH 10, a run voltage of 20 kV, and capillary temperature of 20 C led to complete separation of As(V) and phosphate peaks. Results of these investigations together with recovery test data suggest that separation of the As species from soil solutions can be achieved in less than 5 min with detection limits of 0.50, 0.10, and 0.10 mg L{sup {minus}1} for As(III), As(V), and DMA, respectively.

  16. Determination of ethyl sulfate in human serum and urine by capillary zone electrophoresis.

    Science.gov (United States)

    Jung, Balthasar; Caslavska, Jitka; Thormann, Wolfgang

    2008-10-03

    The use of capillary zone electrophoresis (CZE) with indirect absorbance detection for the analysis of ethyl sulfate (EtS) in serum and urine was investigated. EtS is a direct metabolite of ethanol employed as marker for recent alcohol consumption. Fused-silica capillaries of 60 cm total length were either coated with cetyltrimethylammonium bromide (CTAB, 50 microm I.D. capillary) or poly(diallyldimethylammonium chloride) (PDADMAC, 100 microm I.D. capillary) to allow CZE analyses to be performed with reversed polarity. At pH 2.2 with a maleic acid/phthalic acid background electrolyte, both approaches provided reliable EtS serum levels down to 0.2 mg L(-1) (1.6 microM) for the analysis of solid-phase extracts that were prepared after chloride precipitation. Analysis of urines diluted to a conductivity of 5 S m(-1) and analyzed in the two capillary formats resulted in limits of quantification (LOQs) of 2 and 1 mg L(-1), respectively. With urines adjusted to 10 S m(-1) via dilution or condensation, an LOQ of 0.6 mg L(-1) (4.8 microM) was obtained in the CTAB coated capillary whereas in the PDADMAC-coated capillary of equal length not all matrix components were resolved from EtS. The developed assays are robust and suitable to monitor EtS in samples of individuals who consumed as little as one standard drink of an alcoholic beverage containing about 14 g of ethanol.

  17. Raman spectroscopy and capillary electrophoresis applied to forensic colour inkjet printer inks analysis.

    Science.gov (United States)

    Król, Małgorzata; Karoly, Agnes; Kościelniak, Paweł

    2014-09-01

    Forensic laboratories are increasingly engaged in the examination of fraudulent documents, and what is important, in many cases these are inkjet-printed documents. That is why systematic approaches to inkjet printer inks comparison and identification have been carried out by both non-destructive and destructive methods. In this study, micro-Raman spectroscopy and capillary electrophoresis (CE) were applied to the analysis of colour inkjet printer inks. Micro-Raman spectroscopy was used to study the chemical composition of colour inks in situ on a paper surface. It helps to characterize and differentiate inkjet inks, and can be used to create a spectra database of inks taken from different cartridge brands and cartridge numbers. Capillary electrophoresis in micellar electrophoretic capillary chromatography mode was applied to separate colour and colourless components of inks, enabling group identification of those components which occur in a sufficient concentration (giving intensive peaks). Finally, on the basis of the obtained results, differentiation of the analysed inks was performed. Twenty-three samples of inkjet printer inks were examined and the discriminating power (DP) values for both presented methods were established in the routine work of experts during the result interpretation step. DP was found to be 94.0% (Raman) and 95.6% (CE) when all the analysed ink samples were taken into account, and it was 96.7% (Raman) and 98.4% (CE), when only cartridges with different index numbers were considered. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  18. Modification of gel architecture and TBE/TAE buffer composition to minimize heating during agarose gel electrophoresis.

    Science.gov (United States)

    Sanderson, Brian A; Araki, Naoko; Lilley, Jennifer L; Guerrero, Gilberto; Lewis, L Kevin

    2014-06-01

    Agarose gel electrophoresis of DNA and RNA is routinely performed using buffers containing either Tris, acetate, and EDTA (TAE) or Tris, borate, and EDTA (TBE). Gels are run at a low, constant voltage (∼10 V/cm) to minimize current and asymmetric heating effects, which can induce band artifacts and poor resolution. In this study, alterations of gel structure and conductive media composition were analyzed to identify factors causing higher electrical currents during horizontal slab gel electrophoresis. Current was reduced when thinner gels and smaller chamber buffer volumes were used, but was not influenced by agarose concentration or the presence of ethidium bromide. Current was strongly dependent on the amount and type of EDTA used and on the concentrations of the major acid-base components of each buffer. Interestingly, resolution and the mobilities of circular versus linear plasmid DNAs were also affected by the chemical form and amount of EDTA. With appropriate modifications to gel structure and buffer constituents, electrophoresis could be performed at high voltages (20-25 V/cm), reducing run times by up to 3-fold. The most striking improvements were observed with small DNAs and RNAs (10-100 bp): high voltages and short run times produced sharper bands and higher resolution. Copyright © 2014 Elsevier Inc. All rights reserved.

  19. Gel-eletroforese no diagnóstico da varíola Gel-electrophoresis in the smallpox diagnosis

    Directory of Open Access Journals (Sweden)

    Julio A. Mesquita

    1972-01-01

    Full Text Available O emprego de gel-eletroforese no diagnóstico da varíola, demonstrou ser ao menos trinta vezes (30X mais sensível que o teste de agar-gel, nas condições descritas (tabela I. Doze (12 espécimes, cujos testes convencionais de inoculação em ovos embrionados e de agar-gel resultaram positivos, foram testados em suas diluições originais congeladas por mais de um ano, sendo seis deles revelados por gel-eletroforese enquanto nenhum o foi por agar-gel (tabela II. Trinta e três (33 amostras isoladas no laboratório, foram testadas com material colhido de membrana cório-alantóica da primeira inoculação para o diagnóstico, conservado em glicerina 50%, resultando 15 positivas em gel-eletroforese e apenas 3 em agar-gel (tabela II. Os últimos 60 espécimes recebidos para diagnóstico, através a Campanha de Erradicação da Varíola, também resultaram negativos em gel-eletroforese, que não mostrou falsos-positivos nas condições descritas.The test of gel-electrophoresis applied to the pox virus group showed to be at least thirth times (30X more sensitive than agar-gel test on the described conditions (Table I. Twelve specimens, which were positives form Smallpox in the conventional tests of egg inoculation and agar-gel difusion test, have been screened in their original dilutions frozen for more than 1 year and six of them were still detectable by gel-eletrophoresis, while by agar-gel test any of them was positive (Table II. Thirty three Smallpox isolates have been tested with material from first egg inoculation (chorioallantoic membranes which have been stored in glycerin 50%, at - 15ºC. Fifteen of them were still positive by gel-electrophoresis and only 3 by agar-gel (Table II. The last 60 specimens received for diagnosis from Smallpox Erradication Campaign (CEV, were negatives by both tests. The gel-electrophoresis, did not show false-positives on described conditions.

  20. Sequence-based separation of single-stranded DNA using nucleotides in capillary electrophoresis: focus on phosphate.

    Science.gov (United States)

    Zhang, Xueru; McGown, Linda B

    2013-06-01

    DNA analysis has widespread applicability in biology, medicine, biotechnology, and forensics. DNA separation by length is readily achieved using sieving gels in electrophoresis. Separation by sequence is less simple, generally requiring adequate differences in native or induced conformation or differences in thermal or chemical stability of the strands that are hybridized prior to measurement. We previously demonstrated separation of four single-stranded DNA 76-mers that differ by only a few A-G substitutions based solely on sequence using guanosine-5'-monophosphate (GMP) in the running buffer. We attributed separation to the unique self-assembly of GMP to form higher order structures. Here, we examine an expanded set of 76-mers designed to probe the mechanism of the separation and effects of experimental conditions. We were surprised to find that other ribonucleotides achieved the similar separation to GMP, and that some separation was achieved using sodium phosphate instead of GMP. Potassium phosphate achieved almost as good separations as the ribonucleotides. This suggests that the separation medium provides a physicochemical environment for the DNA that effects strand migration in a sequence-selective manner. Further investigation is needed to determine whether the mechanism involves specific interactions between the phosphates and the DNA strands or is a result of other properties of the separation medium. Phosphate generally has been avoided in DNA separations by capillary gel electrophoresis because its high ionic strength exacerbates Joule heating. Our results suggest that phosphate compounds should be examined for separation of DNA based on sequence. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Northern blots: capillary transfer of RNA from agarose gels and filter hybridization using standard stringency conditions.

    Science.gov (United States)

    Rio, Donald C

    2015-03-02

    In this protocol, an RNA sample, fractionated by gel electrophoresis, is transferred from the gel onto a membrane by capillary transfer. Short-wave UV light is used to fix the transferred RNA to the membrane. The membrane is then pretreated to block nonspecific probe-binding sites, and hybridization of the immobilized RNA to a (32)P-labeled DNA or RNA probe specific for the mRNA of interest is performed. Finally, the membrane is washed and subjected to autoradiography or phosphorimaging. Because exposure to UV cross-links the RNA to the membrane, the membrane can be stripped and hybridized with other probes. The procedure is suitable for detecting poly(A)(+)-selected mRNA or mRNA in total cellular RNA if the target transcript is relatively abundant. Using DNA or RNA probes labeled to 1 × 10(8)-10 × 10(8) cpm/µg, it should be possible to detect ∼5 pg of a specific RNA. © 2015 Cold Spring Harbor Laboratory Press.

  2. Detection of human DNA polymorphisms with a simplified denaturing gradient gel electrophoresis technique

    International Nuclear Information System (INIS)

    Noll, W.W.; Collins, M.

    1987-01-01

    Single base pair differences between otherwise identical DNA molecules can result in altered melting behavior detectable by denaturing gradient gel electrophoresis. The authors have developed a simplified procedure for using denaturing gradient gel electrophoresis to detect base pair changes in genomic DNA. Genomic DNA is digested with restriction enzymes and hybridized in solution to labeled single-stranded probe DNA. The excess probe is then hybridized to complementary phage M13 template DNA, and the reaction mixture is electrophoresed on a denaturing gradient gel. Only the genomic DNA probe hybrids migrate into the gel. Differences in hybrid mobility on the gel indicate base pair changes in the genomic DNA. They have used this technique to identify two polymorphic sites within a 1.2-kilobase region of human chromosome 20. This approach should greatly facilitate the identification of DNA polymorphisms useful for gene linkage studies and the diagnosis of genetic diseases

  3. Interfacing capillary electrophoresis with inductively coupled plasma mass spectrometry for redox speciation of plutonium

    International Nuclear Information System (INIS)

    Ambard, C.; Delorme, A.; Baglan, N.; Aupiais, J.; Pointurier, F.; Madic, C.

    2005-01-01

    A robust and efficient interface between a capilary electrophoresis (CE) and an ICP-MS for actinide speciation studies was developed. This interface was made of two stainless steel T-shape pieces connected to the ICP-MS through a PFA-50 nebulizer. Fast separations (typically in less than 15 min) were obtained. The performances of the technique in terms of chemical separations carried out by the capillary electrophoresis and in terms of detection limits were investigated. Concerning the detection limit of the CE-ICP-MS system for plutonium, it was determined as 5 x 10 -10 mol L -1 or 9 x 10 -18 mol under our injection conditions. The coupling enables to separate at least three (III, V and VI) of the four plutonium oxidation states which can exist in aqueous solutions and to monitor oxidation and reduction processes. (orig.)

  4. 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.

  5. 12-Channel Peltier array temperature control unit for single molecule enzymology studies using capillary electrophoresis.

    Science.gov (United States)

    Craig, Douglas B; Reinfelds, Gundars; Henderson, Anna

    2014-08-01

    Capillary electrophoresis has been used to demonstrate that individual molecules of a given enzyme support different catalytic rates. In order to determine how rate varies with temperature, and determine activation energies for individual β-galactosidase molecules, a 12-channel Peltier array temperature control device was constructed where the temperature of each cell was separately controlled. This array was used to control the temperature of the central 30 cm of a 50 cm long capillary, producing a temperature gradient along its length. Continuous flow single β-galactosidase molecule assays were performed allowing measurement of the catalytic rates at different temperatures. Arrhenius plots were produced and the distribution of activation energies for individual β-galactosidase molecules was found to be 56 ± 10 kJ/mol with a range of 34-72 kJ/mol. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. Determination of Betaine in Forsythia Suspensa by High Performance Capillary Electrophoresis

    Science.gov (United States)

    Liu, Haixing; Dong, Guoliang; Wang, Lintong

    2017-12-01

    This paper presents the determination of betaine content of Forsythia suspensa by high performance capillary electrophoresis (HPCE) method. The borax solution was chosen as buffer solution, and its concentration was 40 mmol with capillary column (75μm×52/60cm) at a constant voltage of 20kV and injecting pressure time of 10s at 20°C. Linearity was kept in the concent ration range of 0.0113-1.45mg·ml-1 of betaine with correlation coefficient of 0.999. The recovery was in the range of 97%-117% (n=5), The content of betaine was 281.5 mg·g-1and RSD value of 9.6% (n=6) in Forsythia suspensa. This method has the advantage of rapid, accurate and good repeatability in separation and determination of betaine in Forsythia suspensa.

  7. Determination of acid-base dissociation constants of azahelicenes by capillary zone electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Ehala, Sille; Míšek, Jiří; Stará, Irena G.; Starý, Ivo; Kašička, Václav

    2008-01-01

    Roč. 31, č. 14 (2008), s. 2686-2693 ISSN 1615-9306 R&D Projects: GA ČR(CZ) GA203/06/1044; GA ČR GA203/08/1428; GA ČR GA203/07/1664; GA MŠk LC512 Institutional research plan: CEZ:AV0Z40550506 Keywords : acidity constant * azahelicenes * non-aqueous capillary electrophoresis Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 2.746, year: 2008

  8. Flavonoids biosynthesis in plants and its further analysis by capillary electrophoresis.

    Science.gov (United States)

    Singh, Baljinder; Kumar, Ashwini; Malik, Ashok Kumar

    2017-03-01

    Flavonoids represent an important bioactive component in plants. Accumulation of flavonoids often occurs in plants subjected to abiotic stresses, including the adaptation of plants to the environment and in overcoming their stress conditions. This fact makes their analysis and determination an attractive field in food science since they can give interesting information on the quality and safety of foods. In this study, we discuss reports on plants flavonoids biosynthesis against abiotic stresses and advances in analytical capillary electrophoresis used for their identification and quantification in plants. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Resolution of a configurationally stable [5]helquat. enantiocomposition analysis of a helicene congener by capillary electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Severa, Lukáš; Koval, Dušan; Novotná, P.; Ončák, M.; Sázelová, Petra; Šaman, David; Slavíček, P.; Urbanová, M.; Kašička, Václav; Teplý, Filip

    2010-01-01

    Roč. 34, č. 6 (2010), s. 1063-1067 ISSN 1144-0546 R&D Projects: GA ČR GA203/09/1614; GA ČR GA203/09/0705; GA ČR(CZ) GA203/08/1428; GA ČR GAP207/10/2391 Institutional research plan: CEZ:AV0Z40550506 Keywords : [5]helquat * capillary electrophoresis * resolution via diastereoisomers Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 2.631, year: 2010

  10. Analysis of ecstasy tablets using capillary electrophoresis with capacitively coupled contactless conductivity detection.

    Science.gov (United States)

    Porto, Suely K S S; Nogueira, Thiago; Blanes, Lucas; Doble, Philip; Sabino, Bruno D; do Lago, Claudimir L; Angnes, Lúcio

    2014-11-01

    A method for the identification of 3,4-methylenedioxymethamphetamine (MDMA) and meta-chlorophenylpiperazine (mCPP) was developed employing capillary electrophoresis (CE) with capacitively coupled contactless conductivity detection (C(4) D). Sample extraction, separation, and detection of "Ecstasy" tablets were performed in fenproporex, caffeine, lidocaine, and cocaine. Separation was performed in <90 sec. The advantages of using C(4) D instead of traditional CE-UV methods for in-field analysis are also discussed. © 2014 American Academy of Forensic Sciences.

  11. Inhibition study of alanine aminotransferase enzyme using sequential online capillary electrophoresis analysis.

    Science.gov (United States)

    Liu, Lina; Chen, Yuanfang; Yang, Li

    2014-12-15

    We report the study of several inhibitors on alanine aminotransferase (ALT) enzyme using sequential online capillary electrophoresis (CE) assay. Using metal ions (Na(+) and Mg(2+)) as example inhibitors, we show that evolution of the ALT inhibition reaction can be achieved by automatically and simultaneously monitoring the substrate consumption and product formation as a function of reaction time. The inhibition mechanism and kinetic constants of ALT inhibition with succinic acid and two traditional Chinese medicines were derived from the sequential online CE assay. Our study could provide valuable information about the inhibition reactions of ALT enzyme. Copyright © 2014 Elsevier Inc. All rights reserved.

  12. Automated dual capillary electrophoresis system with hydrodynamic injection for the concurrent determination of cations and anions

    Energy Technology Data Exchange (ETDEWEB)

    Pham, Thi Thanh Thuy; Mai, Thanh Duc [University of Basel, Department of Chemistry, Spitalstrasse 51, Basel 4056 (Switzerland); Centre for Environmental Technology and Sustainable Development (CETASD), Hanoi University of Science, Nguyen Trai Street 334, Hanoi (Viet Nam); Nguyen, Thanh Dam [Centre for Environmental Technology and Sustainable Development (CETASD), Hanoi University of Science, Nguyen Trai Street 334, Hanoi (Viet Nam); Sáiz, Jorge [Department of Analytical Chemistry, Physical Chemistry and Chemical Engineering – University of Alcalá, Ctra. Madrid-Barcelona km 33.6, Alcalá de Henares, Madrid 28871 (Spain); Pham, Hung Viet, E-mail: phamhungviet@hus.edu.vn [Centre for Environmental Technology and Sustainable Development (CETASD), Hanoi University of Science, Nguyen Trai Street 334, Hanoi (Viet Nam); Hauser, Peter C., E-mail: Peter.Hauser@unibas.ch [University of Basel, Department of Chemistry, Spitalstrasse 51, Basel 4056 (Switzerland)

    2014-09-02

    Highlights: • Concurrent determination of cations and anions was carried out by electrophoretic separation. • Optimized conditions for each class of analystes was possible by using separate capillaries. • Simultaneous hydrodynamic injection was carried out. • Pneumatic actuation was used for flushing and sample handling. • The denitrification of drinking water was successfully demonstrated. - Abstract: The capillary electrophoresis instrument developed for the concurrent determination of cations and anions features two separate capillaries and individual detectors to allow independent optimization for each group of ions. The capillaries are joined in a common injector block. The sample is drawn into the injector with a small membrane pump and automated simultaneous injection into both capillaries is achieved by pressurization of the fluid with compressed air. Flushing of the injector and of the capillaries with the background electrolyte is also carried out automatically by the same means. The buffer consisted of 12 mM histidine and 2 mM 18-crown-6 adjusted to pH 4 with acetic acid and was suitable for the contactless conductivity detection employed. The system was optimized for the determination of cationic NH{sub 4}{sup +} and anionic NO{sub 3}{sup −} and NO{sub 2}{sup −}, and linear calibration curves from about 20 μM up to about 1.5 mM were obtained for these ions. In a test run over 8 h, the reproducibility for the peak areas was within ±7%. For demonstration, the instrument was successfully applied to the concurrent monitoring of the concentrations of the three ions during the biological removal of ammonium from contaminated groundwater in a sequencing batch reactor, where NO{sub 3}{sup −} and NO{sub 2}{sup −} are formed as intermediate products.

  13. Simultaneous determination of rifabutin and human serum albumin in pharmaceutical formulations by capillary electrophoresis.

    Science.gov (United States)

    Ermolenko, Yu; Anshakova, A; Osipova, N; Kamentsev, M; Maksimenko, O; Balabanyan, V; Gelperina, S

    Capillary zone electrophoresis (CZE) was used for determination of rifabutin (RFB), an anti-tuberculosis antibiotic drug, in various pharmaceutical formulations. Apart from that, simultaneous determination of RFB and human serum albumin (HSA) was performed. Electrophoretic behaviour of RFB was examined at various pH levels. CE conditions: a quartz capillary tube (internal diameter 75mm, effective length 50cm, total length 60cm), the capillary temperature was 25°С, the voltage applied to the capillary tube was +20kV, the UV detection wavelength was 214nm, hydrodynamic injection of the sample was performed at 30mbar for 5s, tetraborate buffer solution (0.01М, рН9.2). The obtained results are characterized by high efficiency (number of theoretical plates up to 260,000) and sufficient sensitivity (LOQ starting from 0.02μg/ml for RFB). The obtained data are in good accord with both HPLC results (for RFB) and spectrophotometry (for HSA). Copyright © 2017 Elsevier Inc. All rights reserved.

  14. Use of quasi-isoelectric buffers to limit protein adsorption in capillary zone electrophoresis.

    Science.gov (United States)

    Poitevin, Martine; Hammad, Karim; Ayed, Ichraf; Righetti, Pier Giorgio; Peltre, Gabriel; Descroix, Stephanie

    2008-08-01

    The use of quasi-isoelectric buffers consisting of narrow pH cuts of carrier ampholytes (NC) has been investigated to limit protein adsorption on capillary walls during capillary zone electrophoresis experiments. To quantify protein adsorption on the silica surface, a method derived from that of Towns and Regnier has been developed. alpha-Lactalbumin (14 kDa, pI 4.8) and alpha-chymotrypsinogen A (25 kDa, pI 9.2) have been used as model proteins. Acidic narrow pH cuts of carrier ampholytes (NC, pH 3.0) obtained from fractionation of Serva 4-9 carrier ampholytes were used as BGE in bare-silica capillaries, and allowed to decrease significantly protein adsorption, as compared to experiments performed with classical formate buffer. The use of NC as BGE appeared to be as efficient as the use of polydimethylacrylamide coating to prevent protein adsorption. This increase of protein recovery when using NC was attributed to the interaction of carrier ampholytes with the silica surface, leading to a shielding of the capillary wall.

  15. Capillary electrophoresis of heparin and other glycosaminoglycans using a polyamine running electrolyte

    International Nuclear Information System (INIS)

    Loegel, Thomas N.; Trombley, John D.; Taylor, Richard T.; Danielson, Neil D.

    2012-01-01

    Highlights: ► Ethylenediamine is likely acting as an ion-pairing agent. ► Oversulfated chondroitin sulfate is last peak instead of first peak. ► There is about a factor of five improved detectability with a 12.5 min analysis time. ► Use of a 50 μm ID capillary is possible. - Abstract: This study involves the use of polyamines as potential resolving agents for the capillary electrophoresis (CE) of glycosaminoglycans (GAGs), specifically heparin, dermatan sulfate, chondroitin sulfate, over-sulfated chondroitin sulfate (OSCS), and hyaluronan. All of the compounds can be separated from each other with the exception of chondroitin sulfate and hyaluronan. Using optimization software, the final run conditions are found to be 200 mM ethylenediamine and 45.5 mM phosphate as the electrolyte with −14 V applied across a 50 μm ID × 24.5 cm fused silica capillary at 15 °C. The ion migration order, with OSCS as the last instead of the first peak, is in contrast to previous reports using either a high molarity TRIS or lithium phosphate run buffer with narrower bore capillaries. Total analysis time is 12. 5 min and the relative standard deviation of the heparin migration time is about 2.5% (n = 5). The interaction mechanism between selected polyamines and heparin is explored using conductivity measurements in addition to CE experiments to show that an ion-pairing mechanism is likely.

  16. Alkaline gel electrophoresis assay to detect DNA strand breaks and repair mechanisms in Escherichia coli

    International Nuclear Information System (INIS)

    Mattos, Jose Carlos Pelielo de; Motta, Ellen Serri da; Oliveira, Marcia Betania Nunes de; Dantas, Flavio Jose da Silva; Araujo, Adriano Caldeira de

    2008-01-01

    Reactive oxygen species (ROS) can induce lesions in different cellular targets, including DNA. Stannous chloride (SnCl 2 ) is a ROS generator, leading to lethality in Escherichia coli (E. coli), with the base excision repair (BER) mechanism playing a role in this process. Many techniques have been developed to detect genotoxicity, as comet assay, in eukaryotic cells, and plasmid DNA agarose gel electrophoresis. In this study, an adaptation of the alkaline gel electrophoresis method was carried out to ascertain the induction of strand breaks by SnCl 2 in bacterial DNA, from E. coli BER mutants, and its repair pathway. Results obtained show that SnCl 2 was able to induce DNA strand breaks in all strains tested. Moreover, endonuclease IV and exonuclease III play a role in DNA repair. On the whole, data has shown that the alkaline gel electrophoresis assay could be used both for studying DNA strand breaks induction and for associated repair mechanisms. (author)

  17. Technical improvement to prevent DNA degradation of Leptospira spp. in pulsed field gel electrophoresis.

    Science.gov (United States)

    Ribeiro, R L; Machry, L; Brazil, J M V; Ramos, T M V; Avelar, K E S; Pereira, M M

    2009-08-01

    Leptospirosis is a public health problem. Infection with pathogenic Leptospira occurs by exposure to many environments and is traditionally associated with occupational risk activities. Pulsed-field gel electrophoresis was used to investigate the epidemiological relatedness among Leptospira isolates. However, analysis by PFGE yielded inconclusive data as a result of extensive DNA degradation. This degradation can be significantly reduced by the inclusion of thiourea in the electrophoresis buffer, improving the analysis of DNA banding patterns.

  18. Separation of plant hormones from biofertilizer by capillary electrophoresis using a capillary coated dynamically with polycationic polymers.

    Science.gov (United States)

    Jiang, Ting-Fu; Lv, Zhi-Hua; Wang, Yuan-Hong; Yue, Mei-E

    2006-06-01

    A new, simple and rapid capillary electrophoresis (CE) method, using hexadimethrine bromide (HDB) as electroosmotic flow (EOF) modifier, was developed for the identification and quantitative determination of four plant hormones, including gibberellin A3 (GA3), indole-3-acetic acid (IAA), alpha-naphthaleneacetic acid (NAA) and 4-chlorophenoxyacetic acid (4-CA). The optimum separation was achieved with 20 mM borate buffer at pH 10.00 containing 0.005% (w/v) of HDB. The applied voltage was -25 kV and the capillary temperature was kept constant at 25 degrees C. Salicylic acid was used as internal standard for quantification. The calibration dependencies exhibited good linearity within the ratios of the concentrations of standard samples and internal standard and the ratios of the peak areas of samples and internal standard. The correlation coefficients were from 0.9952 to 0.9997. The relative standard deviations of migration times and peak areas were biofertilizer were successfully determined within 7 min, with satisfactory repeatability and recovery.

  19. Automatic DNA Diagnosis for 1D Gel Electrophoresis Images using Bio-image Processing Technique.

    Science.gov (United States)

    Intarapanich, Apichart; Kaewkamnerd, Saowaluck; Shaw, Philip J; Ukosakit, Kittipat; Tragoonrung, Somvong; Tongsima, Sissades

    2015-01-01

    DNA gel electrophoresis is a molecular biology technique for separating different sizes of DNA fragments. Applications of DNA gel electrophoresis include DNA fingerprinting (genetic diagnosis), size estimation of DNA, and DNA separation for Southern blotting. Accurate interpretation of DNA banding patterns from electrophoretic images can be laborious and error prone when a large number of bands are interrogated manually. Although many bio-imaging techniques have been proposed, none of them can fully automate the typing of DNA owing to the complexities of migration patterns typically obtained. We developed an image-processing tool that automatically calls genotypes from DNA gel electrophoresis images. The image processing workflow comprises three main steps: 1) lane segmentation, 2) extraction of DNA bands and 3) band genotyping classification. The tool was originally intended to facilitate large-scale genotyping analysis of sugarcane cultivars. We tested the proposed tool on 10 gel images (433 cultivars) obtained from polyacrylamide gel electrophoresis (PAGE) of PCR amplicons for detecting intron length polymorphisms (ILP) on one locus of the sugarcanes. These gel images demonstrated many challenges in automated lane/band segmentation in image processing including lane distortion, band deformity, high degree of noise in the background, and bands that are very close together (doublets). Using the proposed bio-imaging workflow, lanes and DNA bands contained within are properly segmented, even for adjacent bands with aberrant migration that cannot be separated by conventional techniques. The software, called GELect, automatically performs genotype calling on each lane by comparing with an all-banding reference, which was created by clustering the existing bands into the non-redundant set of reference bands. The automated genotype calling results were verified by independent manual typing by molecular biologists. This work presents an automated genotyping tool from DNA

  20. Automatic DNA Diagnosis for 1D Gel Electrophoresis Images using Bio-image Processing Technique

    Science.gov (United States)

    2015-01-01

    Background DNA gel electrophoresis is a molecular biology technique for separating different sizes of DNA fragments. Applications of DNA gel electrophoresis include DNA fingerprinting (genetic diagnosis), size estimation of DNA, and DNA separation for Southern blotting. Accurate interpretation of DNA banding patterns from electrophoretic images can be laborious and error prone when a large number of bands are interrogated manually. Although many bio-imaging techniques have been proposed, none of them can fully automate the typing of DNA owing to the complexities of migration patterns typically obtained. Results We developed an image-processing tool that automatically calls genotypes from DNA gel electrophoresis images. The image processing workflow comprises three main steps: 1) lane segmentation, 2) extraction of DNA bands and 3) band genotyping classification. The tool was originally intended to facilitate large-scale genotyping analysis of sugarcane cultivars. We tested the proposed tool on 10 gel images (433 cultivars) obtained from polyacrylamide gel electrophoresis (PAGE) of PCR amplicons for detecting intron length polymorphisms (ILP) on one locus of the sugarcanes. These gel images demonstrated many challenges in automated lane/band segmentation in image processing including lane distortion, band deformity, high degree of noise in the background, and bands that are very close together (doublets). Using the proposed bio-imaging workflow, lanes and DNA bands contained within are properly segmented, even for adjacent bands with aberrant migration that cannot be separated by conventional techniques. The software, called GELect, automatically performs genotype calling on each lane by comparing with an all-banding reference, which was created by clustering the existing bands into the non-redundant set of reference bands. The automated genotype calling results were verified by independent manual typing by molecular biologists. Conclusions This work presents an

  1. Immunochemical characterization of Mycobacterium leprae antigens by the SDS-polyacrylamide gel electrophoresis immunoperoxidase technique (SGIP) using patients' sera

    NARCIS (Netherlands)

    Klatser, P. R.; van Rens, M. M.; Eggelte, T. A.

    1984-01-01

    In this study the SDS-polyacrylamide gel electrophoresis immunoperoxidase (SGIP) assay was used for characterizing the antigenic components of Mycobacterium leprae using patients' sera. This technique involved the separation of mycobacterial sonicates on SDS-polyacrylamide gels, longitudinal

  2. Separation and determination of flavonoids in three traditional chinese medicines by capillary electrophoresis with amperometric detection.

    Science.gov (United States)

    Wang, Wei; Lin, Ping; Ma, Lihong; Xu, Kaixuan; Lin, Xiuli

    2016-04-01

    Flavonoids are important active ingredients in many traditional Chinese medicines. In this paper, capillary electrophoresis with amperometric detection was employed to separate and detect eight flavonoids, rutin, quercetrin, quercetin, kaempferol, kaempferide, catechin, apigenin, and luteolin, in a home-made capillary electrophoresis device. Under the separation voltage of 2000 V, the eight flavonoids could be completely separated within 33 min in 18 mM borax running buffer at pH 10.2. Good linear relationships were obtained for all analytes and the detection limits for flavonoids ranged from 0.46 to 0.85 μM. Then, the method was applied to separate and determine the flavonoids in three traditional Chinese medicines, hippophae rhamnoides, hypericum perforatum, and cacumen platycladi. Finally, rutin, kaempferol, quercetin, and quercetrin were discovered in these medicines and the concentrations ranged from 0.28 to 9.94 mg/g. The recoveries of flavonoids ranged from 84.7 to 113%, which showed the high reliability of this method. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Capillary electrophoresis to determine entrapment efficiency of a nanostructured lipid carrier loaded with piroxicam

    Directory of Open Access Journals (Sweden)

    Jessica Otarola

    2015-02-01

    Full Text Available A simple and fast capillary electrophoresis method has been developed to determine the amount of piroxicam loaded in a drug delivery system based on nanostructured lipid carriers (NLCs. The entrapment efficiency of the nanostructured lipid carrier was estimated by measuring the concentration of drug not entrapped in a suspension of NLC. The influence of different parameters on migration times, peak symmetry, efficiency and resolution was studied; these parameters included the pH of the electrophoretic buffer solution and the applied voltage. The piroxicam peak was obtained with a satisfactory resolution. The separation was carried out using a running buffer composed of 50 mM ammonium acetate and 13.75 mM ammonia at pH 9. The optimal voltage was 20 kV and the cartridge temperature was 20 °C. The corresponding calibration curve was linear over the range of 2.7–5.4 µg/mL of NLC suspension. The reproducibility of migration time and peak area were investigated, and the obtained RSD% values (n=5 were 0.99 and 2.13, respectively. Keywords: Capillary electrophoresis, Drug delivery system, Nanostructured lipid carrier, Piroxicam

  4. An accessible micro-capillary electrophoresis device using surface-tension-driven flow

    Science.gov (United States)

    Mohanty, Swomitra K.; Warrick, Jay; Gorski, Jack; Beebe, David J.

    2010-01-01

    We present a rapidly fabricated micro-capillary electrophoresis chip that utilizes surface-tension-driven flow for sample injection and extraction of DNA. Surface-tension-driven flow (i.e. passive pumping) injects a fixed volume of sample that can be predicted mathematically. Passive pumping eliminates the need for tubing, valves, syringe pumps, and other equipment typically needed for interfacing with microelectrophoresis chips. This method requires a standard micropipette to load samples before separation, and remove the resulting bands after analysis. The device was made using liquid phase photopolymerization to rapidly fabricate the chip without the need of special equipment typically associated with the construction of microelectrophoresis chips (e.g. cleanroom). Batch fabrication time for the device presented here was 1.5 h including channel coating time to suppress electroosmotic flow. Devices were constructed out of poly-isobornyl acrylate and glass. A standard microscope with a UV source was used for sample detection. Separations were demonstrated using Promega BenchTop 100 bp ladder in hydroxyl ethyl cellulose (HEC) and oligonucleotides of 91 and 118 bp were used to characterize sample injection and extraction of DNA bands. The end result was an inexpensive micro-capillary electrophoresis device that uses tools (e.g. micropipette, electrophoretic power supplies, and microscopes) already present in most labs for sample manipulation and detection, making it more accessible for potential end users. PMID:19425002

  5. On-line detection of small radioactive ions by capillary electrophoresis

    International Nuclear Information System (INIS)

    Klunder, G.L.; Andrews, J.E. Jr.; Russo, R.E.

    1994-01-01

    Worldwide environmental interests have placed a great demand on developing techniques for rapid characterization of contaminated soil and groundwater. Detection of radioactive contaminants is necessary for monitoring effluents from nuclear processes or to assure proper long term storage of radioactive waste. The authors have been investigating the chemistry required to separate representative radioactive small cations and anions by capillary electrophoresis. In order to evaluate the separation chemistry, detection of stable isotopes of the representative ions was achieved by indirect absorption for cations and direct absorption for anions. Several buffer systems which have been considered in the optimization of the separations will be discussed. The authors have designed and tested two on-line radioactivity detectors for capillary electrophoresis. An on-line solid state CdTe detector was constructed for this study and a scintillation detector has been designed using a high gain photodiode light sensor. Different scintillation materials have been tested. Comparison of the detectors, design considerations, efficiency and limits of detection will be presented

  6. ANALYSIS OF GLYCANS DERIVED FROM GLYCOCONJUGATES BY CAPILLARY ELECTROPHORESIS-MASS SPECTROMETRY

    Science.gov (United States)

    Mechref, Yehia

    2012-01-01

    The high structural variation of glycan derived from glycoconjugates, which substantially increases with the molecular size of a protein, contributes to the complexity of glycosylation patterns commonly associated with glycoconjugates. In the case of glycoproteins, such variation originates from the multiple glycosylation sites of proteins and the number of glycan structures associated with each site (microheterogeneity). The ability to comprehensively characterize highly complex mixture of glycans has been analytically stimulating and challenging. Although the most powerful mass spectrometric (MS) and tandem MS techniques are capable of providing a wealth of structural information, they are still not able to readily identify isomeric glycan structures without high order tandem MS (MSn). The analysis of isomeric glycan structures has been attained using several separation methods, including high-pH anion exchange chromatography (HPAEC), hydrophilic interaction chromatography (HILIC) and gas chromatography (GC). However, capillary electrophoresis (CE) and microfluidics capillary electrophoresis (MCE) offer high separation efficiency and resolutions, allowing the separation of closely related glycan structures. Therefore, interfacing CE and MCE to MS is a powerful analytical approach, allowing potentially comprehensive and sensitive analysis of complex glycan samples. This review describes and discusses the utility of different CE and MCE approaches in the structural characterization of glycoproteins and the feasibility of interfacing these approaches to mass spectrometry. PMID:22180203

  7. Mobility-based correction for accurate determination of binding constants by capillary electrophoresis-frontal analysis.

    Science.gov (United States)

    Qian, Cheng; Kovalchik, Kevin A; MacLennan, Matthew S; Huang, Xiaohua; Chen, David D Y

    2017-06-01

    Capillary electrophoresis frontal analysis (CE-FA) can be used to determine binding affinity of molecular interactions. However, its current data processing method mandate specific requirement on the mobilities of the binding pair in order to obtain accurate binding constants. This work shows that significant errors are resulted when the mobilities of the interacting species do not meet these requirements. Therefore, the applicability of CE-FA in many real word applications becomes questionable. An electrophoretic mobility-based correction method is developed in this work based on the flux of each species. A simulation program and a pair of model compounds are used to verify the new equations and evaluate the effectiveness of this method. Ibuprofen and hydroxypropyl-β-cyclodextrinare used to demonstrate the differences in the obtained binding constant by CE-FA when different calculation methods are used, and the results are compared with those obtained by affinity capillary electrophoresis (ACE). The results suggest that CE-FA, with the mobility-based correction method, can be a generally applicable method for a much wider range of applications. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Simultaneous determination of phenylethanoid glycosides and aglycones by capillary zone electrophoresis with running buffer modifier.

    Science.gov (United States)

    Dong, Shuqing; Gao, Ruibin; Yang, Yan; Guo, Mei; Ni, Jingman; Zhao, Liang

    2014-03-15

    Although the separation efficiency of capillary electrophoresis (CE) is much higher than that of other chromatographic methods, it is sometimes difficult to adequately separate the complex ingredients in biological samples. This article describes how one effective and simple way to develop the separation efficiency in CE is to add some modifiers to the running buffer. The suitable running buffer modifier β-cyclodextrin (β-CD) was explored to fast and completely separate four phenylethanoid glycosides and aglycones (homovanillyl alcohol, hydroxytyrosol, 3,4-dimethoxycinnamic acid, and caffeic acid) in Lamiophlomis rotata (Lr) and Cistanche by capillary zone electrophoresis with ultraviolet (UV) detection. It was found that when β-CD was used as running buffer modifier, a baseline separation of the four analytes could be accomplished in less than 20 min and the detection limits were as low as 10(-3) mg L(-1). Other factors affecting the CE separation, such as working potential, pH value and ionic strength of running buffer, separation voltage, and sample injection time, were investigated extensively. Under the optimal conditions, a successful practical application on the determination of Lr and Cistanche samples confirmed the validity and practicability of this method. Copyright © 2014 Elsevier Inc. All rights reserved.

  9. Simultaneous determination of flavonoids in chrysanthemum by capillary zone electrophoresis with running buffer modifiers.

    Science.gov (United States)

    Zhang, Shan; Dong, Shuqing; Chi, Langzhu; He, Pingang; Wang, Qingjiang; Fang, Yuzhi

    2008-08-15

    Despite the separation efficiency of capillary electrophoresis (CE) is much higher than other chromatographic methods, it is sometimes difficult to perfectly separate the complex ingredients in biological samples. One possible and simple way to develop the separation effect in CE is to add some modifiers in the running buffer. In this paper, the suitable running buffer modifiers were explored to simultaneously separate and detect six typical flavonoids (apigenin, luteolin, kaempferol, quercetin, (+)-catechin and (-)-epicatechin) which are the main active ingredients in chrysanthemum by capillary zone electrophoresis with amperometric detection (CZE-AD). It was found that when beta-cyclodextrin (beta-CD) and the mixture of methanol and ethanol were used as running buffer modifiers, a baseline separation of the six analytes could be accomplished in less than 20 min and the detection limits were as low as 10(-7) or 10(-8)gm l(-1). Other factors affecting the CZE separation, such as working potential, pH value and ionic strength of running buffer, separation voltage and sample injection time were extensively investigated. Under the optimum conditions, a successful practical application on the determination of chrysanthemum samples confirmed the validity and practicability of this method.

  10. Ocular Proteomics with Emphasis on Two-Dimensional Gel Electrophoresis and Mass Spectrometry

    Directory of Open Access Journals (Sweden)

    Honoré Bent

    2010-01-01

    Full Text Available Abstract The intention of this review is to provide an overview of current methodologies employed in the rapidly developing field of ocular proteomics with emphasis on sample preparation, two-dimensional polyacrylamide gel electrophoresis (2D-PAGE and mass spectrometry (MS. Appropriate sample preparation for the diverse range of cells and tissues of the eye is essential to ensure reliable results. Current methods of protein staining for 2D-PAGE, protein labelling for two-dimensional difference gel electrophoresis, gel-based expression analysis and protein identification by MS are summarised. The uses of gel-free MS-based strategies (MuDPIT, iTRAQ, ICAT and SILAC are also discussed. Proteomic technologies promise to shed new light onto ocular disease processes that could lead to the discovery of strong novel biomarkers and therapeutic targets useful in many ophthalmic conditions.

  11. Qualitative and quantitative analysis of cations and anions using ion selective detectors in capillary electrophoresis

    International Nuclear Information System (INIS)

    Nann, A.

    1994-01-01

    The present work reports on the application of ion-selective microelectrodes as potentiometric detectors for the qualitative and quantitative analysis of cations and anions separated by capillary electrophoresis. Due to the high internal resistance of microelectrodes, their potentials are strongly affected by external electrical fields. Therefore, the influence of the electrophoretic field on the electrode response had to be kept at a minimum. With the electrode tip inserted in the capillary aperture (on-column detection), heavy drifts and noise of the signals were observed, mainly because the electrophoretic potential is superimposed on the Nernstian electrode response. As the potential inside the capillary is site-dependent, already minor movements and vibrations not perceptible under the light microscopy cause unacceptable disturbances of the electrode signal. One possibility to solve the problem consists in post- or off-column detection, i.e., with the detector located outside the influence of the electrophoretic field. If quantitative analyses with maximum resolution are to be achieved, only on-column detection is suitable because outside the capillary, the separation efficiency drops drastically. By etching the detector-side capillary end to a conical aperture, the field strength in the last 10 μm fell approximately 1/25 as compared with that in a cylindrical one. Thus, potential drifts and noise were reduced correspondingly so that on-column detection can also be used for potentiometric detection. To obtain quantitative results, the signals of the ion-selective detector were first delogarithmized and then integrated over time. Thus, it was possible to quantify cations and anions with a coefficient of variations ≤5%. (author) figs., tabs., 179 refs

  12. Cyclodextrine Screening for the Chiral Separation of Amlodipine Enantiomers by Capillary Electrophoresis

    Directory of Open Access Journals (Sweden)

    Gabriel Hancu

    2015-03-01

    Full Text Available Purpose: Amlodipine is a long acting, dihydropyridine type calcium channel blocker frequently used in the treatment of hypertension and coronary insufficiency. The calcium channel blocking activity resides primarily in the S-amlodipine enantiomer, while R-amlodipine is a potent inhibitor of smooth muscle cell migration. Methods: In this study capillary electrophoresis was applied for the enantiomeric separation of amlodipine using different native and derivatized; neutral and charged cyclodextrines as chiral selectors. The effects of pH and composition of the background electrolyte, concentration and type of chiral selector, capillary temperature, running voltage and injection parameters have been investigated. Results: Stereoselective interactions were observed when using α-CD, β-CD, HP-β-CD, RAMEB, CM-β-CD and SBE-β-CD. Optimized separation conditions consisted on a 50 mM phosphate buffer, pH – 3.0, 20 mM RAMEB as chiral selector, + 25 kV applied voltage, 15°C temperature and UV detection at 238 nm. Using the optimized electrophoretic conditions we succeeded the chiral separation of amlodipine enantiomers in approximately 6 minute, the order of migration being R-amlodipine followed by S-amlodipine. The method was successfully applied for the determination of amlodipine enantiomers from commercially available pharmaceuticals. The linearity range, limits of detection and quantification, precision and accuracy were determined and the results obtained confirmed that the method was suitable for this purpose. Conclusion: It can be concluded that the proposed capillary electrophoresis methods can be useful for routine pharmaceutical applications with benefits of its effectivity, simplicity, short analysis time and low consumption of analytes, solvents and chiral selectors.

  13. Affinity in electrophoresis.

    Science.gov (United States)

    Heegaard, Niels H H

    2009-06-01

    The journal Electrophoresis has greatly influenced my approaches to biomolecular affinity studies. The methods that I have chosen as my main tools to study interacting biomolecules--native gel and later capillary zone electrophoresis--have been the topic of numerous articles in Electrophoresis. Below, the role of the journal in the development and dissemination of these techniques and applications reviewed. Many exhaustive reviews on affinity electrophoresis and affinity CE have been published in the last few years and are not in any way replaced by the present deliberations that are focused on papers published by the journal.

  14. Heparin/heparan sulfate analysis by covalently modified reverse polarity capillary zone electrophoresis-mass spectrometry.

    Science.gov (United States)

    Sanderson, Patience; Stickney, Morgan; Leach, Franklin E; Xia, Qiangwei; Yu, Yanlei; Zhang, Fuming; Linhardt, Robert J; Amster, I Jonathan

    2018-04-13

    Reverse polarity capillary zone electrophoresis coupled to negative ion mode mass spectrometry (CZE-MS) is shown to be an effective and sensitive tool for the analysis of glycosaminoglycan mixtures. Covalent modification of the inner wall of the separation capillary with neutral or cationic reagents produces a stable and durable surface that provides reproducible separations. By combining CZE-MS with a cation-coated capillary and a sheath flow interface, a rapid and reliable method has been developed for the analysis of sulfated oligosaccharides from dp4 to dp12. Several different mixtures have been separated and detected by mass spectrometry. The mixtures were selected to test the capability of this approach to resolve subtle differences in structure, such as sulfation position and epimeric variation of the uronic acid. The system was applied to a complex mixture of heparin/heparan sulfate oligosaccharides varying in chain length from dp3 to dp12 and more than 80 molecular compositions were identified by accurate mass measurement. Copyright © 2018 Elsevier B.V. All rights reserved.

  15. Analysis of compositional monosaccharides in fungus polysaccharides by capillary zone electrophoresis.

    Science.gov (United States)

    Hu, Yuanyuan; Wang, Tong; Yang, Xingbin; Zhao, Yan

    2014-02-15

    A rapid analytical method of capillary zone electrophoresis (CZE) was established for the simultaneous separation and determination of 10 monosaccharides (aldoses and uronic acids). The monosaccharides were labeled with 1-phenyl-3-methyl-5-pyrazolone (PMP), and subsequently separated using an uncoated capillary (50 μm i.d. × 58.5 cm) and detected by UV at 245 nm with pH 11.0, 175 mM borate buffer at voltage 20 kV and capillary temperature 25 °C by CZE. The 10 PMP-labeled monosaccharides were rapidly baseline separated within 20 min. The optimized CZE method was successfully applied to the simultaneous separation and identification of the monosaccharide composition in Termitomyces albuminosus polysaccharides (TAPs) and Panus giganteus polysaccharides (PGPs). The quantitative recovery of the component monosaccharides in the fungus polysaccharides was in the range of 92.0-101.0% and the CV value was lower than 3.5%. The results demonstrate that the proposed CZE method is precise and practical for the monosaccharide analysis of fungus polysaccharides. Copyright © 2013 Elsevier Ltd. All rights reserved.

  16. Quantitative Determination of Lercanidipine Enantiomers in Commercial Formulations by Capillary Electrophoresis

    Directory of Open Access Journals (Sweden)

    Luciana Pereira Lourenço

    2015-01-01

    Full Text Available An enantioselective method based on capillary electrophoresis (CE using cyclodextrin (CD as chiral selector was developed and validated for determination of lercanidipine (LER enantiomers, a drug calcium channel blocker which exerts antihypertensive effects of long duration, in a pharmaceutical formulation. Optimum separation of LER enantiomers was obtained on a 50 cm × 50 μm id capillary using a sodium acetate buffer solution 200 mmol/L pH 4.0 containing 10 mmol/L of 2,3,6-o-methyl-β-cyclodextrin (TM-β-CD as background electrolyte. The capillary temperature and voltage were 15°C and 25 kV, respectively, hydrodynamic injection and detection at 237 nm. Linearity was obtained in the range 12.5–100 μg/mL for both enantiomers (r≥0.995. The RSD (% and relative errors (E, % obtained in precision and accuracy studies (intraday and interday were lower than 5%. After validation, the method was applied to quantify the enantiomers of LER in commercial tablets and the results were satisfactory in terms of accuracy and precision, both less than 5%. Therefore, this method was found to be appropriate for enantioselective quality control of LER enantiomers in pharmaceutical formulations.

  17. pI-Control in comparative fluorescence gel electrophoresis (CoFGE) using amphoteric azo dyes

    Czech Academy of Sciences Publication Activity Database

    Hanneken, M.; Šlais, Karel; König, S.

    2015-01-01

    Roč. 3, JUN (2015), s. 221-228 ISSN 2352-3409 Institutional support: RVO:68081715 Keywords : 2D-PAGE * CoFGE * Gel electrophoresis Subject RIV: CB - Analytical Chemistry, Separation http://www.sciencedirect.com/science/article/pii/S2352340915000414

  18. New Approach for Segmentation and Quantification of Two-Dimensional Gel Electrophoresis Images

    DEFF Research Database (Denmark)

    Anjo, Antonio dos; Laurell Blom Møller, Anders; Ersbøll, Bjarne Kjær

    2011-01-01

    Motivation: Detection of protein spots in two-dimensional gel electrophoresis images (2-DE) is a very complex task and current approaches addressing this problem still suffer from significant shortcomings. When quantifying a spot, most of the current software applications include a lot of backgro...

  19. Detection of hydrolysis of lipid post-translational modifications during gel-electrophoresis-based proteomic protocol

    Czech Academy of Sciences Publication Activity Database

    Žídková, Jitka; Řehulka, Pavel; Chmelík, Josef

    2007-01-01

    Roč. 7, č. 15 (2007), s. 2507-2510 ISSN 1615-9853 R&D Projects: GA MŠk 1M0570 Institutional research plan: CEZ:AV0Z40310501 Keywords : ester hydrolysis * gel electrophoresis * MALDI-TOF MS Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 5.479, year: 2007

  20. Separation of hydrolytically active components of cellulase from Myrothecium verrucaria by starch gel electrophoresis

    NARCIS (Netherlands)

    Ritter, F.J.; Prins-van der Meulen, P.Y.F.; Marel, T. van der

    1968-01-01

    Using starch gel electrophoresis according to Smithies, desalted crude cellulase from Myrothecium verrucqria was separated into at least 12 protein zones. These were tested on their activity towards p-nitrophenyl-β-D-glucoside, sodium carboxymethylcellulose and α-cellulose. They were all

  1. Comparison of Pseudomonas aeruginosa isolates from mink by serotyping and pulsed-field gel electrophoresis

    DEFF Research Database (Denmark)

    Hammer, Anne Sofie; Pedersen, Karl; Andersen, Thomas Holmen

    2003-01-01

    Isolates of Pseudomonas aeruginosa from clinical infections in mink were subjected to serotyping and pulsed-field gel electrophoresis (PFGE) using SpeI. A total of 212 isolates of P aeruginosa from the year 1998 to 2001 were included in this study: 168 isolates from mink obtained from 74 farm out...

  2. Pulsed-field Gel Electrophoresis for Salmonella Infection Surveillance, Texas, USA, 2007

    Centers for Disease Control (CDC) Podcasts

    This podcast describes monitoring of the use of pulsed-field gel electrophoresis for Salmonella surveillance in Houston, Texas. CDC microbiologist Peter Gerner-Smidt discusses the importance of the PulseNet national database in surveillance of food-borne infections.

  3. A Novel Strategy for Characterization of Glycosylated Proteins Separated by Gel Electrophoresis

    DEFF Research Database (Denmark)

    Larsen, Martin; Skottrup, Peter; Enghild, Jan J.

    2005-01-01

    . We present a new technique, which allows full characterization of low amounts of glycoproteins separated by gel electrophoresis. The method takes advantage of sequential specific and non-specific enzymatic treatment, followed by selective purification and characterization of the glycopeptides using...

  4. Introducing Proteomics in the Undergraduate Curriculum: A Simple 2D Gel Electrophoresis Exercise with Serum Proteins

    Science.gov (United States)

    Kim, Thomas D.; Craig, Paul A.

    2010-01-01

    Two-dimensional gel electrophoresis (2DGE) remains an important tool in the study of biological systems by proteomics. While the use of 2DGE is commonplace in research publications, there are few instructional laboratories that address the use of 2DGE for analyzing complex protein samples. One reason for this lack is the fact that the preparation…

  5. Progress in two-dimensional polyacrylamide gel electrophoresis and application in radiation research

    International Nuclear Information System (INIS)

    Wang Zhidong; Chen Xiaohua

    2003-01-01

    Two-dimensional polyacrylamide gel electrophoresis is the key separation technique in proteomics research, which is designed by protein character: molecular weight and PI. Some progress has been made in disease mechanism detection, tumor indicator research and drug development. This technique also has some potential application in radiation research

  6. Investigation of two different anoxia models by 2-dimensional gel electrophoresis

    DEFF Research Database (Denmark)

    Wulff, Tune; Jessen, Flemming; Hoffmann, Else Kay

    2006-01-01

    anoxia obtained by NaN3 is a widely used model for simulating anoxia (Ossum et al., 2004). The effects of anoxia were studied by protein expression analysis using 2-dimensional gel electrophoresis followed by MS/MS. In this way we were able to separate more than 1500 protein spots with an apparent range...

  7. Improvement of fragment and primer selection for mutation detection by denaturing gradient gel electrophoresis

    NARCIS (Netherlands)

    Wu, Y; Hayes, VM; Osinga, J; Mulder, IM; Looman, MWG; Buys, CHCM; Hofstra, RMW

    1998-01-01

    Denaturing gradient gel electrophoresis (DGGE) is one of the most powerful methods for mutation detection currently available. For successful application the appropriate selection of PCR fragments and PCR primers is crucial. The sequence of interest should always be within the domain with the lowest

  8. Analysis of rRNA gene methylation in Arabidopsis thaliana by CHEF-Conventional 2D gel electrophoresis

    Science.gov (United States)

    Mohannath, Gireesha; Pikaard, Craig S.

    2017-01-01

    Summary Contour-clamped homogenous electric field (CHEF) gel electrophoresis, a variant of Pulsed-field gel electrophoresis (PFGE), is a powerful technique for resolving large fragments of DNA (10 kb to 9 Mb). CHEF has many applications including the physical mapping of chromosomes, artificial chromosomes and sub-chromosomal DNA fragments, etc. Here we describe the use of CHEF and two-dimensional gel electrophoresis to analyze rRNA gene methylation patterns within the two ~ 4 million base pair nucleolus organizer regions (NORs) of Arabidopsis thaliana. The method involves CHEF gel electrophoresis of agarose-embedded DNA following restriction endonuclease digestion to cut the NORs into large but resolvable segments, followed by digestion with methylation-sensitive restriction endonucleases and conventional (or CHEF) gel electrophoresis, in a second dimension. Resulting products are then detected by Southern blotting or PCR analyses capable of discriminating rRNA gene subtypes. PMID:27576719

  9. Serum protein concentrations from clinically healthy horses determined by agarose gel electrophoresis.

    Science.gov (United States)

    Riond, Barbara; Wenger-Riggenbach, Bettina; Hofmann-Lehmann, Regina; Lutz, Hans

    2009-03-01

    Serum protein electrophoresis is a useful screening test in equine laboratory medicine. The method can provide valuable information about changes in the concentrations of albumin and alpha-, beta-, and gamma-globulins and thereby help characterize dysproteinemias in equine patients. Reference values for horses using agarose gel as a support medium have not been reported. The purpose of this study was to establish reference intervals for serum protein concentrations in adult horses using agarose gel electrophoresis and to assess differences between warm-blooded and heavy draught horses. In addition, the precision of electrophoresis for determining fraction percentages and the detection limit were determined. Blood samples were obtained from 126 clinically healthy horses, including 105 Thoroughbreds and 21 heavy draught horses of both sexes and ranging from 2 to 20 years of age. The total protein concentration was determined by an automated biuret method. Serum protein electrophoresis was performed using a semi-automated agarose gel electrophoresis system. Coefficients of variation (CVs) were calculated for within-run and within-assay precision. Data from warm-blooded and draught horses were compared using the Mann-Whitney U test. Within-run and within-assay CVs were draught horses and so combined reference intervals (2.5-97.5%) were calculated for total protein (51.0-72.0 g/L), albumin (29.6-38.5 g/L), alpha(1)-globulin (1.9-3.1 g/L), alpha(2)-globulin (5.3-8.7 g/L), beta(1)-globulin (2.8-7.3g/L), beta(2)-globulin (2.2-6.0 g/L), and gamma-globulin (5.8-12.7 g/L) concentrations, and albumin/globulin ratio (0.93-1.65). Using agarose gel as the supporting matrix for serum protein electrophoresis in horses resulted in excellent resolution and accurate results that facilitated standardization into 6 protein fractions.

  10. Analysis of Proteins, Protein Complexes, and Organellar Proteomes Using Sheathless Capillary Zone Electrophoresis - Native Mass Spectrometry

    Science.gov (United States)

    Belov, Arseniy M.; Viner, Rosa; Santos, Marcia R.; Horn, David M.; Bern, Marshall; Karger, Barry L.; Ivanov, Alexander R.

    2017-12-01

    Native mass spectrometry (MS) is a rapidly advancing field in the analysis of proteins, protein complexes, and macromolecular species of various types. The majority of native MS experiments reported to-date has been conducted using direct infusion of purified analytes into a mass spectrometer. In this study, capillary zone electrophoresis (CZE) was coupled online to Orbitrap mass spectrometers using a commercial sheathless interface to enable high-performance separation, identification, and structural characterization of limited amounts of purified proteins and protein complexes, the latter with preserved non-covalent associations under native conditions. The performance of both bare-fused silica and polyacrylamide-coated capillaries was assessed using mixtures of protein standards known to form non-covalent protein-protein and protein-ligand complexes. High-efficiency separation of native complexes is demonstrated using both capillary types, while the polyacrylamide neutral-coated capillary showed better reproducibility and higher efficiency for more complex samples. The platform was then evaluated for the determination of monoclonal antibody aggregation and for analysis of proteomes of limited complexity using a ribosomal isolate from E. coli. Native CZE-MS, using accurate single stage and tandem-MS measurements, enabled identification of proteoforms and non-covalent complexes at femtomole levels. This study demonstrates that native CZE-MS can serve as an orthogonal and complementary technique to conventional native MS methodologies with the advantages of low sample consumption, minimal sample processing and losses, and high throughput and sensitivity. This study presents a novel platform for analysis of ribosomes and other macromolecular complexes and organelles, with the potential for discovery of novel structural features defining cellular phenotypes (e.g., specialized ribosomes). [Figure not available: see fulltext.

  11. Capillary electrophoresis of heparin and other glycosaminoglycans using a polyamine running electrolyte

    Energy Technology Data Exchange (ETDEWEB)

    Loegel, Thomas N.; Trombley, John D.; Taylor, Richard T. [Department of Chemistry and Biochemistry, Miami University, Oxford, OH 45056 (United States); Danielson, Neil D., E-mail: danielnd@muohio.edu [Department of Chemistry and Biochemistry, Miami University, Oxford, OH 45056 (United States)

    2012-11-13

    Highlights: Black-Right-Pointing-Pointer Ethylenediamine is likely acting as an ion-pairing agent. Black-Right-Pointing-Pointer Oversulfated chondroitin sulfate is last peak instead of first peak. Black-Right-Pointing-Pointer There is about a factor of five improved detectability with a 12.5 min analysis time. Black-Right-Pointing-Pointer Use of a 50 {mu}m ID capillary is possible. - Abstract: This study involves the use of polyamines as potential resolving agents for the capillary electrophoresis (CE) of glycosaminoglycans (GAGs), specifically heparin, dermatan sulfate, chondroitin sulfate, over-sulfated chondroitin sulfate (OSCS), and hyaluronan. All of the compounds can be separated from each other with the exception of chondroitin sulfate and hyaluronan. Using optimization software, the final run conditions are found to be 200 mM ethylenediamine and 45.5 mM phosphate as the electrolyte with -14 V applied across a 50 {mu}m ID Multiplication-Sign 24.5 cm fused silica capillary at 15 Degree-Sign C. The ion migration order, with OSCS as the last instead of the first peak, is in contrast to previous reports using either a high molarity TRIS or lithium phosphate run buffer with narrower bore capillaries. Total analysis time is 12. 5 min and the relative standard deviation of the heparin migration time is about 2.5% (n = 5). The interaction mechanism between selected polyamines and heparin is explored using conductivity measurements in addition to CE experiments to show that an ion-pairing mechanism is likely.

  12. High-Throughput Genetic Analysis and Combinatorial Chiral Separations Based on Capillary Electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Zhong, Wenwan [Iowa State Univ., Ames, IA (United States)

    2003-01-01

    Capillary electrophoresis (CE) offers many advantages over conventional analytical methods, such as speed, simplicity, high resolution, low cost, and small sample consumption, especially for the separation of enantiomers. However, chiral method developments still can be time consuming and tedious. They designed a comprehensive enantioseparation protocol employing neutral and sulfated cyclodextrins as chiral selectors for common basic, neutral, and acidic compounds with a 96-capillary array system. By using only four judiciously chosen separation buffers, successful enantioseparations were achieved for 49 out of 54 test compounds spanning a large variety of pKs and structures. Therefore, unknown compounds can be screened in this manner to identify optimal enantioselective conditions in just one rn. In addition to superior separation efficiency for small molecules, CE is also the most powerful technique for DNA separations. Using the same multiplexed capillary system with UV absorption detection, the sequence of a short DNA template can be acquired without any dye-labels. Two internal standards were utilized to adjust the migration time variations among capillaries, so that the four electropherograms for the A, T, C, G Sanger reactions can be aligned and base calling can be completed with a high level of confidence. the CE separation of DNA can be applied to study differential gene expression as well. Combined with pattern recognition techniques, small variations among electropherograms obtained by the separation of cDNA fragments produced from the total RNA samples of different human tissues can be revealed. These variations reflect the differences in total RNA expression among tissues. Thus, this Ce-based approach can serve as an alternative to the DNA array techniques in gene expression analysis.

  13. Fractionation of SWNT/nucleic acid complexes by agarose gel electrophoresis

    International Nuclear Information System (INIS)

    Vetcher, Alexandre A; Srinivasan, Srimeenakshi; Vetcher, Ivan A; Abramov, Semen M; Kozlov, Mikhail; Baughman, Ray H; Levene, Stephen D

    2006-01-01

    We show that aqueous dispersions of single-walled carbon nanotubes (SWNTs), prepared with the aid of nucleic acids (NAs) such as RNA or DNA, can be separated into fractions using agarose gel electrophoresis. In a DC electric field, SWNT/NA complexes migrate in the gel in the direction of positive potential to form well-defined bands. Raman spectroscopy as a function of band position shows that nanotubes having different spectroscopic properties possess different electrophoretic mobilities. The migration patterns for SWNT/RNA and SWNT/DNA complexes differ. Parallel elution of the SWNT/NA complexes from the gel during electrophoresis and subsequent characterization by AFM reveals differences in nanotube diameter, length and curvature. The results suggest that fractionation of nanotubes can be achieved by this procedure. We discuss factors affecting the mobility of the nanotube complexes and propose analytical applications of this technique

  14. Fractionation of SWNT/nucleic acid complexes by agarose gel electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Vetcher, Alexandre A [Institute of Biomedical Sciences and Technology and Department of Molecular and Cell Biology, University of Texas at Dallas, Richardson, TX 75083 (United States); Srinivasan, Srimeenakshi [Institute of Biomedical Sciences and Technology and Department of Molecular and Cell Biology, University of Texas at Dallas, Richardson, TX 75083 (United States); Vetcher, Ivan A [Institute of Biomedical Sciences and Technology and Department of Molecular and Cell Biology, University of Texas at Dallas, Richardson, TX 75083 (United States); Abramov, Semen M [NanoTech Institute, University of Texas at Dallas, Richardson, TX 75083 (United States); Kozlov, Mikhail [NanoTech Institute, University of Texas at Dallas, Richardson, TX 75083 (United States); Baughman, Ray H [NanoTech Institute, University of Texas at Dallas, Richardson, TX 75083 (United States); Levene, Stephen D [Institute of Biomedical Sciences and Technology and Department of Molecular and Cell Biology, University of Texas at Dallas, Richardson, TX 75083 (United States)

    2006-08-28

    We show that aqueous dispersions of single-walled carbon nanotubes (SWNTs), prepared with the aid of nucleic acids (NAs) such as RNA or DNA, can be separated into fractions using agarose gel electrophoresis. In a DC electric field, SWNT/NA complexes migrate in the gel in the direction of positive potential to form well-defined bands. Raman spectroscopy as a function of band position shows that nanotubes having different spectroscopic properties possess different electrophoretic mobilities. The migration patterns for SWNT/RNA and SWNT/DNA complexes differ. Parallel elution of the SWNT/NA complexes from the gel during electrophoresis and subsequent characterization by AFM reveals differences in nanotube diameter, length and curvature. The results suggest that fractionation of nanotubes can be achieved by this procedure. We discuss factors affecting the mobility of the nanotube complexes and propose analytical applications of this technique.

  15. Optimization of Large Gel 2D Electrophoresis for Proteomic Studies of Skeletal Muscle

    Science.gov (United States)

    Reed, Patrick W.; Densmore, Allison; Bloch, Robert J.

    2013-01-01

    We describe improved methods for large format, 2-dimensional gel electrophoresis (2-DE) that improve protein solubility and recovery, minimize proteolysis, and reduce the loss of resolution due to contaminants and manipulations of the gels, and thus enhance quantitative analysis of protein spots. Key modifications are: (i) the use of 7M urea + 2 M thiourea, instead of 9M urea, in sample preparation and in the tops of the gel tubes; (ii) standardized deionization of all solutions containing urea with a mixed bed ion exchange resin and removal of urea from the electrode solutions; and (iii) use of a new gel tank and cooling device that eliminate the need to run two separating gels in the SDS dimension. These changes make 2D-GE analysis more reproducible and sensitive, with minimal artifacts. Application of this method to the soluble fraction of muscle tissues reliably resolves ~1800 protein spots in adult human skeletal muscle and over 2800 spots in myotubes. PMID:22589104

  16. Rapid capillary electrophoresis approach for the quantification of ewe milk adulteration with cow milk.

    Science.gov (United States)

    Trimboli, Francesca; Morittu, Valeria Maria; Cicino, Caterina; Palmieri, Camillo; Britti, Domenico

    2017-10-13

    The substitution of ewe milk with more economic cow milk is a common fraud. Here we present a capillary electrophoresis method for the quantification of ewe milk in ovine/bovine milk mixtures, which allows for the rapid and inexpensive recognition of ewe milk adulteration with cow milk. We utilized a routine CE method for human blood and urine proteins analysis, which fulfilled the separation of skimmed milk proteins in alkaline buffer. Under this condition, ovine and bovine milk exhibited a recognizable and distinct CE protein profiles, with a specific ewe peak showing a reproducible migration zone in ovine/bovine mixtures. Based on ewe specific CE peak, we developed a method for ewe milk quantification in ovine/bovine skimmed milk mixtures, which showed good linearity, precision and accuracy, and a minimum amount of detectable fraudulent cow milk equal to 5%. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. Capillary Electrophoresis Single-Strand Conformational Polymorphisms as a Method to Differentiate Algal Species

    Directory of Open Access Journals (Sweden)

    Alice Jernigan

    2015-01-01

    Full Text Available Capillary electrophoresis single-strand conformational polymorphism (CE-SSCP was explored as a fast and inexpensive method to differentiate both prokaryotic (blue-green and eukaryotic (green and brown algae. A selection of two blue-green algae (Nostoc muscorum and Anabaena inaequalis, five green algae (Chlorella vulgaris, Oedogonium foveolatum, Mougeotia sp., Scenedesmus quadricauda, and Ulothrix fimbriata, and one brown algae (Ectocarpus sp. were examined and CE-SSCP electropherogram “fingerprints” were compared to each other for two variable regions of either the 16S or 18S rDNA gene. The electropherogram patterns were remarkably stable and consistent for each particular species. The patterns were unique to each species, although some common features were observed between the different types of algae. CE-SSCP could be a useful method for monitoring changes in an algae species over time as potential shifts in species occurred.

  18. Detection of Elevated Signaling Amino Acids in Human Diabetic Vitreous by Rapid Capillary Electrophoresis

    Directory of Open Access Journals (Sweden)

    Miao-Jen Lu

    2007-01-01

    Full Text Available Elevated glutamate is implicated in the pathology of PDR. The ability to rapidly assess the glutamate and amino acid content of vitreous provides a more complete picture of the chemical changes occurring at the diabetic retina and may lead to a better understanding of the pathology of PDR. Vitreous humor was collected following vitrectomies of patients with PDR and control conditions of macular hole or epiretinal membrane. A capillary electrophoresis method was developed to quantify glutamate and arginine. The analysis is relatively fast (<6 minutes and utilizes a poly(ethyleneoxide and sodium dodecylsulfate run buffer. Both amino acid levels show significant increases in PDR patients versus controls and are comparable to other reports. The levels of vitreal glutamate vary inversely with the degree of observed hemorrhage. The results demonstrate a rapid method for assessment of a number of amino acids to characterize the chemical changes at the diabetic retina to better understand tissue changes and potentially identify new treatments.

  19. Sensitive determination of phenolic acids in extra-virgin olive oil by capillary zone electrophoresis.

    Science.gov (United States)

    Carrasco Pancorbo, Alegría; Cruces-Blanco, Carmen; Segura Carretero, Antonio; Fernández Gutiérrez, Alberto

    2004-11-03

    A sensitive, rapid, efficient, and reliable method for the separation and determination of phenolic acids by capillary zone electrophoresis has been carried out. A detailed method optimization was carried out to separate 14 different compounds by studying parameters such as pH, type and concentration of buffer, applied voltage, and injection time. The separation was performed within 16 min, using a 25 mM sodium borate buffer (pH 9.6) at 25 kV with 8 s of hydrodynamic injection. With this method and using a liquid-liquid extraction system, with recovery values around 95%, it has been possible to detect small quantities of phenolic acids in olive oil samples. This is apparently the first paper showing the quantification of this specific family of phenolic compounds in virgin olive oil samples.

  20. Applications of capillary electrophoresis with chemiluminescence detection in clinical, environmental and food analysis. A review

    International Nuclear Information System (INIS)

    Lara, Francisco J.; Airado-Rodríguez, Diego; Moreno-González, David; Huertas-Pérez, José F.; García-Campaña, Ana M.

    2016-01-01

    This paper reviews the latest developments and analytical applications of chemiluminescence detection coupled to capillary electrophoresis (CE-CL). Different sections considering the most common CL systems have been included, such as the tris(2,2′-bipyridine)ruthenium(II) system, the luminol and acridinium derivative reactions, the peroxyoxalate CL or direct oxidations. Improvements in instrumental designs, new strategies for improving both resolution and sensitivity, and applications in different fields such as clinical, pharmaceutical, environmental and food analysis have been included. This review covers the literature from 2010 to 2015. - Highlights: • An up-to-date critical review about the evolution of CE-CL is presented. • Tris(2,2′-bipyridine)ruthenium(II) and luminol as the most used CL systems. • Instrumental designs and strategies for improving resolution and sensitivity. • Applications in clinical, pharmaceutical, environmental and food analysis.

  1. Protein A Detection Based on Quantum Dots-Antibody Bioprobe Using Fluorescence Coupled Capillary Electrophoresis

    Directory of Open Access Journals (Sweden)

    Lin Qiu

    2014-01-01

    Full Text Available In this report, fluorescence detection coupled capillary electrophoresis (CE-FL was used to detect Protein A. Antibody was first labeled with Cy5 and then mixed with quantum dots (QDs to form QDs-antibody bioprobe. Further, we observed fluorescence resonance energy transfer (FRET from QDs donor to Cy5 acceptor. The bioprobe was formed and brought QDs and Cy5 close enough to allow FRET to occur. After adding protein A, the FRET system was broken and caused the FRET signal to decrease. Thus, a new method for the determination of protein A was proposed based on the FRET signal changes. This study provides a new trail of thought for the detection of protein.

  2. Determination of Inorganic Ion Profiles of Illicit Drugs by Capillary Electrophoresis.

    Science.gov (United States)

    Evans, Elizabeth; Costrino, Carolina; do Lago, Claudimir L; Garcia, Carlos D; Roux, Claude; Blanes, Lucas

    2016-11-01

    A portable capillary electrophoresis instrument with dual capacitively coupled contactless conductivity detection (C 4 D) was used to determine the inorganic ionic profiles of three pharmaceutical samples and precursors of two illicit drugs (contemporary samples of methylone and para-methoxymethamphetamine). The LODs ranged from 0.10 μmol/L to 1.25 μmol/L for the 10 selected cations, and from 0.13 μmol/L to 1.03 μmol/L for the eight selected anions. All separations were performed in less than 6 min with migration times and peak area RSD values ranging from 2 to 7%. The results demonstrate the potential of the analysis of inorganic ionic species to aid in the identification and/or differentiation of unknown tablets, and real samples found in illicit drug manufacture scenarios. From the resulting ionic fingerprint, the unknown tablets and samples can be further classified. © 2016 American Academy of Forensic Sciences.

  3. Enantioseparations of amino acids by capillary array electrophoresis with 532 nm laser induced fluorescence detection.

    Science.gov (United States)

    Liu, Kaiying; Wang, Li

    2013-06-21

    Capillary array electrophoresis (CAE) is a promising technique for multiple enantiomeric separations. Carboxytetramethylrhodamine succinimidyl ester (TAMRA SE), a rhodamine-core fluorescent probe, has rarely been applied as an original precolumn derivatization reagent for chiral amino acid (AA) analysis so far. For these purposes, high-throughput enantiomeric separations of 12 TAMRA SE-AAs by a home-made 532 nm CAE-LIF scanner are presented. The effect of cyclodextrins (CDs) and a variety of organic modifiers was quickly investigated. Baseline separations were achieved in 100 mM Tris-borate buffer (pH 10.0) containing 2 mM β-CD and 10 mM hexamethylenediamine (HDA). Multiple determination of the enantiomeric excess (ee) in non-racemic mixtures of alanine is successfully presented. Copyright © 2013 Elsevier B.V. All rights reserved.

  4. Determination of Betaine in Lycium Barbarum L. by High Performance Capillary Electrophoresis

    Science.gov (United States)

    Liu, Haixing; Wang, Chunyan; Peng, Xuewei

    2017-12-01

    This paper presents the determination of betaine content in Lycium barbarum L. by high performance capillary electrophoresis (HPCE) method. The borax solution was chosen as buffer solution, and its concentration was 40 mmol at a constant voltage of 20kV and injecting pressure time of 10s at 20°C. Linearity was kept in the concent ration range of 0.0113∼1.45mg of betaine with correlation coefficient of 0.9. The recovery was in the range of 97.95%∼126% (n=4). The sample content of betaine was 29.3mg/g and RSD 6.4% (n=6). This method is specific, simple and rapid and accurate, which is suitable for the detection of the content of betaine in Lycium barbarum L.

  5. Determination of recombinant hirudin structural deviants by capillary zone electrophoresis augmented with buffer additives.

    Science.gov (United States)

    Dönges, Reiner; Brazel, Dieter

    2002-12-06

    The polypeptide hirudin is a potent and specific thrombin inhibitor used in anticoagulant therapy and naturally occurring in medicinal leech. Using gene-technology methods, recombinant (r) hirudin can be produced on a large scale. Purity evaluation of the synthesized r-hirudin is essential to monitor co-expressed structural deviants and degradation products before therapeutic use. Although the well established RP-HPLC analysis appears to be the method of choice, in the case of r-hirudin baseline separation of the structural deviants is not necessarily achieved. Capillary zone electrophoresis augmented with buffer additives was used as a complementary technique to separate r-hirudin successfully from several similar species, in order to provide characterization information, as well as performing purity control and stability studies.

  6. Application of 2,3-Naphthalenediamine in Labeling Natural Carbohydrates for Capillary Electrophoresis

    Directory of Open Access Journals (Sweden)

    Jim-Min Fang

    2012-06-01

    Full Text Available Neutral and acidic monosaccharide components in Ganoderma lucidum polysaccharide are readily labeled with 2,3-naphthalenediamine, and the resulting saccharide-naphthimidazole (NAIM derivatives are quantified by capillary electrophoresis (CE in borate buffer. Using sulfated-α-cyclodextrin as the chiral selector, enantiomers of monosaccharide-NAIMs are resolved on CE in phosphate buffer, allowing a simultaneous determination of the absolute configuration and sugar composition in the mucilage polysaccharide of a medicinal herb Dendrobium huoshanense. Together with the specific enzymatic reactions of various glycoside hydrolases on the NAIM derivatives of glycans, the structures of natural glycans can be deduced from the digestion products identified by CE analysis. Though heparin dissachrides could be successfully derived with the NAIM-labeling method, the heparin derivatives with the same degree of sulfation could not be separated by CE.

  7. New cellular automaton designed to simulate geometration in gel electrophoresis

    Science.gov (United States)

    Krawczyk, M. J.; Kułakowski, K.; Maksymowicz, A. Z.

    2002-08-01

    We propose a new kind of cellular automaton to simulate transportation of molecules of DNA through agarose gel. Two processes are taken into account: reptation at strong electric field E, described in the particle model, and geometration, i.e. subsequent hookings and releases of long molecules at and from gel fibres. The automaton rules are deterministic and they are designed to describe both processes within one unified approach. Thermal fluctuations are not taken into account. The number of simultaneous hookings is limited by the molecule length. The features of the automaton are: (i) the size of the cell neighbourhood for the automaton rule varies dynamically, from nearest neighbors to the entire molecule; (ii) the length of the time step is determined at each step according to dynamic rules. Calculations are made up to N=244 reptons in a molecule. Two subsequent stages of the motion are found. Firstly, an initial set of random configurations of molecules is transformed into a more ordered phase, where most molecules are elongated along the applied field direction. After some transient time, the mobility μ reaches a constant value. Then, it varies with N as 1/ N for long molecules. The band dispersion varies with time t approximately as Nt1/2. Our results indicate that the well-known plateau of the mobility μ vs. N does not hold at large electric fields.

  8. Screening of chemokine receptor CCR4 antagonists by capillary zone electrophoresis

    Directory of Open Access Journals (Sweden)

    Zhe Sun

    2011-11-01

    Full Text Available CC chemokine receptor 4 (CCR4 is a kind of G-protein-coupled receptor, which plays a pivotal role in allergic inflammation. The interaction between 2-(2-(4-chloro-phenyl-5-{[(naphthalen-1-ylmethyl-carbamoyl]-methyl}-4-oxo-thiazolidin-3-yl-N-(3-morpholin-4-yl-propyl-acetamide (S009 and the N-terminal extracellular tail (ML40 of CCR4 has been validated to be high affinity by capillary zone electrophoresis (CZE. The S009 is a known CCR4 antagonist. Now, a series of new thiourea derivatives have been synthesized. Compared with positive control S009, they were screened using ML40 as target by CZE to find some new drugs for allergic inflammation diseases. The synthesized compounds XJH-5, XJH-4, XJH-17 and XJH-1 displayed the interaction with ML40, but XJH-9, XJH-10, XJH-11, XJH-12, XJH-13, XJH-14, XJH-3, XJH-8, XJH-6, XJH-7, XJH-15, XJH-16 and XJH-2 did not bind to ML40. Both qualification and quantification characterizations of the binding were determined. The affinity of the four compounds was valued by the binding constant, which was similar with the results of chemotactic experiments. The established CEZ method is capable of sensitive and fast screening for a series of lactam analogs in the drug discovery for allergic inflammation diseases. Keywords: Capillary zone electrophoresis, CCR4 antagonist, 2-(2-(4-chloro-phenyl-5-{[(naphthalen-1-ylmethyl-carbamoyl]-methyl}-4-oxo-thiazolidin-3-yl-N-(3-morpholin-4-yl-propyl-acetamide, Interactions, Structural modification

  9. Simultaneous Detection of 13 Key Bacterial Respiratory Pathogens by Combination of Multiplex PCR and Capillary Electrophoresis.

    Science.gov (United States)

    Jiang, Lu Xi; Ren, Hong Yu; Zhou, Hai Jian; Zhao, Si Hong; Hou, Bo Yan; Yan, Jian Ping; Qin, Tian; Chen, Yu

    2017-08-01

    Lower respiratory tract infections continue to pose a significant threat to human health. It is important to accurately and rapidly detect respiratory bacteria. To compensate for the limits of current respiratory bacteria detection methods, we developed a combination of multiplex polymerase chain reaction (PCR) and capillary electrophoresis (MPCE) assay to detect thirteen bacterial pathogens responsible for lower respiratory tract infections, including Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis, Pseudomonas aeruginosa, Klebsiella pneumoniae, Escherichia coli, Staphylococcus aureus, Mycoplasma pneumoniae, Legionella spp., Bordetella pertussis, Mycobacterium tuberculosis complex, Corynebacterium diphtheriae, and Streptococcus pyogenes. Three multiplex PCR reactions were built, and the products were analyzed by capillary electrophoresis using the high-throughput DNA analyzer. The specificity of the MPCE assay was examined and the detection limit was evaluated using DNA samples from each bacterial strain and the simulative samples of each strain. This assay was further evaluated using 152 clinical specimens and compared with real-time PCR reactions. For this assay, three nested-multiplex-PCRs were used to detect these clinical specimens. The detection limits of the MPCE assay for the 13 pathogens were very low and ranged from 10-7 to 10-2 ng/μL. Furthermore, analysis of the 152 clinical specimens yielded a specificity ranging from 96.5%-100.0%, and a sensitivity of 100.0% for the 13 pathogens. This study revealed that the MPCE assay is a rapid, reliable, and high-throughput method with high specificity and sensitivity. This assay has great potential in the molecular epidemiological survey of respiratory pathogens. Copyright © 2017 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.

  10. Native fluorescence detection of biomolecular and pharmaceutical compounds in capillary electrophoresis: detector designs, performance and applications: A review

    NARCIS (Netherlands)

    de Kort, B.J.; de Jong, G.J.; Somsen, G.W.

    2013-01-01

    This review treats the coupling of capillary electrophoresis (CE) with fluorescence detection (Flu) for the analysis of natively fluorescent biomolecular and pharmaceutical compounds. CE-Flu combines the excellent separation efficiency of CE with the high selectivity and sensitivity of Flu. In

  11. 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.

  12. A chiral capillary electrophoresis method for ropivacaine hydrochloride in pharmaceutical formulations : Validation and comparison with chiral liquid chromatography

    NARCIS (Netherlands)

    Sänger-Van De Griend, C. E.; Wahlström, H.; Gröningsson, K.; Widahl-Näsman, Monica E.

    A capillary electrophoresis method for the determination of the enantiomeric purity of the local anaesthetic ropivacaine hydrochloride in injection solutions has been validated. The method showed the required limit of quantitation of 0.1% enantiomeric impurity. Good performances were shown for

  13. Validation of a capillary electrophoresis method for the enantiomeric purity testing of ropivacaine, a new local anaesthetic compound

    NARCIS (Netherlands)

    Sänger-Van De Griend, C. E.; Gröningsson, K.

    A capillary electrophoresis method for the determination of the enantiomeric purity of ropivacaine, a new local anaesthetic compound developed by Astra Pain Control AB, has been validated. The method showed the required limit of quantitation of 0.1%, enantiomeric impurity and proved to be robust.

  14. Electromembrane extraction of amino acids from body fluids followed by capillary electrophoresis with capacitively coupled contactless conductivity detection

    Czech Academy of Sciences Publication Activity Database

    Strieglerová, Lenka; Kubáň, Pavel; Boček, Petr

    2011-01-01

    Roč. 1218, č. 37 (2011), s. 6248-6255 ISSN 0021-9673 R&D Projects: GA ČR GAP206/10/1219 Institutional research plan: CEZ:AV0Z40310501 Keywords : electromembrane extraction * biological samples * capillary electrophoresis Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 4.531, year: 2011

  15. Ultraviolet-absorbing organic anions in uremic serum separated by capillary zone electrophoresis, and quantification of hippuric acid

    NARCIS (Netherlands)

    Schoots, A.C.; Verheggen, T.P.E.M.; Vries, de P.M.J.M.; Everaerts, F.M.

    1990-01-01

    Organic anions accumulated in blood serum of patients with chronic renal failure were separated by a novel technique: closed-system capillary zone electrophoresis (CZE) in a pH6 carrier-electrolyte system. Hippuric acid (HA), p-hydroxyhippuric acid, and uric acid were identified by their co-elution

  16. Quantification in capillary electrophoresis-mass spectrometry : Long- and short-term variance components and their compensation using internal standards

    NARCIS (Netherlands)

    Ohnesorge, Jens; Sänger-van de Griend, Cari; Wätzig, Hermann

    Different approaches were chosen to examine ionization reproducibility of analytes after separation by capillary electrophoresis-mass spectrometry (CE-MS) in a commercially available sheath-flow electrospray interface. For this task three different standard samples were examined. Sample 1 contained

  17. Determination of phenprocoumon in plasma and urine using at-line solid-phase extraction-capillary electrophoresis.

    NARCIS (Netherlands)

    Veraart, J.R.; Gooijer, C.; Lingeman, H.; Velthorst, N.H.; Brinkman, U.A.T.

    1998-01-01

    The use of capillary electrophoresis (CE) for the analysis of biological samples is rather problematic because of the large number of interferences present in the matrix. One of the possibilities to solve such problems is to couple solid-phase extraction (SPE) at-line with CE, a technique developed

  18. Hair analysis for illicit drugs by using capillary zone electrophoresis-electrospray ionization-ion trap mass spektrometry

    Czech Academy of Sciences Publication Activity Database

    Gottardo, R.; Bortolotti, F.; De Paoli, G.; Pascali, J. P.; Mikšík, Ivan; Tagliaro, F.

    2007-01-01

    Roč. 1159, 1-2 (2007), s. 185-189 ISSN 0021-9673 Institutional research plan: CEZ:AV0Z50110509 Keywords : capillary electrophoresis * hair analysis Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.641, year: 2007

  19. Determination of simple carbohydrates in citrus juices by using capillary electrophoresis with indirect detection and high performance liquid chromatography

    Czech Academy of Sciences Publication Activity Database

    Cabálková, J.; Chmelík, Josef

    96(S), - (2002), s. S150-S152 ISSN 0009-2770. [Meeting of Chemistry & Life /2./. Brno, 10.09.2002-11.09.2002] Institutional research plan: CEZ:AV0Z4031919 Keywords : carbohydrates * citrus juice * capillary electrophoresis Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 0.336, year: 2002

  20. Analysis of ethyl glucuronide in human serum by capillary electrophoresis with sample self-stacking and indirect detection

    Czech Academy of Sciences Publication Activity Database

    Křivánková, Ludmila; Caslavska, J.; Malášková, Hana; Gebauer, Petr; Thormann, W.

    2005-01-01

    Roč. 1081, č. 1 (2005), s. 2-8 ISSN 0021-9673 R&D Projects: GA AV ČR IAA4031401 Institutional research plan: CEZ:AV0Z40310501 Keywords : ethyl glucuronide * capillary electrophoresis * serum Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.096, year: 2005

  1. Affinity capillary electrophoresis and quantum mechanical calculations applied to investigation of [Gly(6)]-antamanide binding with sodium and potassium ions

    Czech Academy of Sciences Publication Activity Database

    Pangavhane, Sachin; Böhm, S.; Makrlík, E.; Ruzza, P.; Kašička, Václav

    2017-01-01

    Roč. 38, č. 12 (2017), s. 1551-1559 ISSN 0173-0835 R&D Projects: GA ČR(CZ) GA15-01948S Institutional support: RVO:61388963 Keywords : affinity capillary electrophoresis * density functional theory * [Gly(6)]-antamanide * peptide complex * stability constant Subject RIV: CB - Analytical Chemistry, Separation OBOR OECD: Analytical chemistry Impact factor: 2.744, year: 2016

  2. PCR/LDR/capillary electrophoresis for detection of single-nucleotide differences between fetal and maternal DNA in maternal plasma.

    Science.gov (United States)

    Yi, Ping; Chen, Zhuqin; Zhao, Yan; Guo, Jianxin; Fu, Huabin; Zhou, Yuanguo; Yu, Lili; Li, Li

    2009-03-01

    The discovery of fetal DNA in maternal plasma has opened up an approach for noninvasive diagnosis. We have now assessed the possibility of detecting single-nucleotide differences between fetal and maternal DNA in maternal plasma by polymerase chain reaction (PCR)/ligase detection reaction((LDR)/capillary electrophoresis. PCR/LDR/capillary electrophoresis was applied to detect the genotype of c.454-397T>gene (ESR1) from experimental DNA models of maternal plasma at different sensitivity levels and 13 maternal plasma samples.alphaC in estrogen receptor. (1) Our results demonstrated that the technique could discriminate low abundance single-nucleotide mutation with a mutant/normal allele ratio up to 1:10 000. (2) Examination of ESR1 c.454-397T>C genotypes by using the method of restriction fragment length analysis was performed in 25 pregnant women, of whom 13 pregnant women had homozygous genotypes. The c.454-397T>C genotypes of paternally inherited fetal DNA in maternal plasma of these 13 women were detected by PCR/LDR/capillary electrophoresis, which were accordant with the results of umbilical cord blood. PCR/LDR/capillary electrophoresis has very high sensitivity to distinguish low abundance single nucleotide differences and can discriminate point mutations and single-nucleotide polymorphisms(SNPs) of paternally inherited fetal DNA in maternal plasma.

  3. Determination of free radical reaction products and metabolites of salicylic acid using capillary electrophoresis and micellar electrokinetic chromatography

    NARCIS (Netherlands)

    Coolen, S.A.J.; Huf, F.A.; Reijenga, J.C.

    1998-01-01

    Hydroxylated radical products of salicylic acid are often used as a relative measurement in free radical research. Several analytical methods exist to determine the amount of 2,3-dihydroxybenzoic acid and 2,5-dihydroxybenzoic acid. In this study we use capillary zone electrophoresis (CZE) and

  4. Performance of a liquid-junction interface for capillary electrophoresis mass spectrometry using continuous-flow fast-atom bombardment

    NARCIS (Netherlands)

    Reinhoud, N.J.; Niessen, W.M.A.; Tjaden, U.R.; Gramberg, L.G.; Verheij, E.R.; Greef, J. van der

    1989-01-01

    The on-line coupling of capillary electrophoresis and mass spectrometry using a continuous-flow fast-atom bombardment system in combination with a liquid-junction interface is described. The influence of the liquid-junction coupling on the efficiency and the resolution is investigated. Qualitative

  5. Fluorescence- and capillary electrophoresis (CE)-based SSR DNA fingerprinting and a molecular identity database for the Louisiana sugarcane industry

    Science.gov (United States)

    A database of Louisiana sugarcane molecular identity has been constructed and is being updated annually using FAM or HEX or NED fluorescence- and capillary electrophoresis (CE)-based microsatellite (SSR) fingerprinting information. The fingerprints are PCR-amplified from leaf DNA samples of current ...

  6. 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

  7. Sensitive determination of malondialdehyde in exhaled breath condensate and biological fluids by capillary electrophoresis with laser induced fluorescence detection

    Czech Academy of Sciences Publication Activity Database

    Lačná, J.; Foret, František; Kubáň, Petr

    2017-01-01

    Roč. 169, JUL (2017), s. 85-90 ISSN 0039-9140 Grant - others:GA ČR(CZ) GA13-21919S Keywords : malondialdehyde * capillary electrophoresis * laser induced fluorescence * blood plasma * saliva Subject RIV: CB - Analytical Chemistry, Separation OBOR OECD: Analytical chemistry Impact factor: 4.162, year: 2016

  8. Structural and conformational variants of human beta2-microglobulin characterized by capillary electrophoresis and complementary separation methods

    DEFF Research Database (Denmark)

    Heegaard, Niels H H; Rovatti, Luca; Nissen, Mogens H

    2003-01-01

    The small (Mr = 11729) serum protein beta2-microglobulin is prone to precipitate as amyloid in a protein conformational disorder (PCD) that occurs in a significant number of patients on chronic hemodialysis. Analyses by capillary electrophoresis (CE) were undertaken to study beta2-microglobulin...

  9. Ultraviolet laser-induced fluorescence detection strategies in capillary electrophoresis: determination of naphthalene sulphonates in river water.

    NARCIS (Netherlands)

    Kok, S.J.; Isberg, I.C.K.; Gooijer, C.; Brinkman, U.A.T.; Velthorst, N.H.

    1998-01-01

    Various UV-laser-induced fluorescence detection strategies for capillary electrophoresis (CE) are compared, i.e. two UV-laser systems (a pulsed laser providing up to 25 mW of tunable emission, applied at 280, 290 and 325 nm, and a continuous wave (cw) laser providing up to 100 mW of 257 nm emission)

  10. Computational simulation of migration and dispersion in free capillary zone electrophoresis, I: Description of the theoretical model

    NARCIS (Netherlands)

    Reijenga, J.C.; Kenndler, E.

    1994-01-01

    An instrument simulator was developed for high-performance capillary electrophoresis which allows for fast graphic illustration of the effect of a large number of variables on the shape of the electropherogram. The input data of the separands are values of pK and mobilities at 25°C and infinite

  11. Influence of ignored and well-known zone distortions on the separation performance of proteins in capillary free zone electrophoresis with special reference to analysis in polyacrylamide-coated fused silica capillaries in various buffers. I. Theoretical studies.

    Science.gov (United States)

    Hjertén, Stellan; Mohabbati, Sheila; Westerlund, Douglas

    2004-10-22

    the peaks become broad. Therefore, different types of buffers should be tested when high resolution is required. The relation sigma2 (the variance of the interaction between a protein and the buffer constituents) = constant x u (the mobility) seems to be valid for all proteins in the applied sample, at least when they have similar molecular masses. To facilitate the understanding of the progress of a free zone electrophoresis experiment, we have discussed in simple terms how the concentrations of the background electrolytes become rearranged during a run and why the difference between the mobilities of the proteins and the mobilities of the background electrolyte determines whether a peak exhibits fronting or tailing. A theoretical analysis of zone broadening in capillary zone electrophoresis, chromatography, and electrochromatography indicates that electrochromatography in homogeneous gels might be the only chromatographic technique which can compete in performance with free electrophoresis. Using an equation, valid not only for electrophoresis, but also for chromatography and centrifugation, the mobility of a concentration boundary has been calculated for the first time and was, as expected, low. Equations based on the Kohlrausch regulating function do not permit such calculations. Another regulating function (the H function) and some of its characteristics are briefly discussed. The theoretical discussions in this paper and the experimental studies in Part II show that high-performance electrophoresis deserves its prefix when the runs are designed to give minimum zone broadening. Some guidelines are given to facilitate this optimization. The plate numbers are so high that the resolution cannot be increased by more than 30% even if they approach the theoretically maximum values.

  12. Comparison of the capillary and agarose electrophoresis based multiple locus VNTR (variable number of tandem repeats) analysis (MLVA) on Mycobacterium bovis isolates.

    Science.gov (United States)

    Jenkins, A O; Venter, E H; Hutamo, K; Godfroid, J

    2010-09-28

    Electrophoretic techniques that can be used for genotyping of bacterial pathogens ranges from manual, low-cost, agarose gels to high-throughput capillary electrophoresis sequencing machines. These two methods are currently employed in the electrophoresis of PCR products used in multiple locus VNTR (variable number of tandem repeats) analysis (MLVA), i.e. the agarose electrophoresis (AE) and the capillary electrophoresis (CE). Some authors have suggested that clusters generated by AE are less reliable than those generated by CE and that the latter is a more sensitive technique than the former when typing Mycobacterium tuberculosis complex (MTC) isolates. Because such a claim could have significant consequences for investigators in this field, a comparison was made on 19 Belgian Mycobacterium bovis strains which had previously been genotyped using CE VNTR analysis. The VNTR profiles of the CE VNTR analysis were compared with those obtained by AE VNTR analysis at 14 VNTR loci. Our results indicated that there were no differences in copy numbers at all loci tested when the copy numbers obtained by the AE VNTR analysis were compared with those obtained by CE VNTR analysis. The use of AE VNTR analysis in mycobacterial genotyping does not alter the sensitivity of the MLVA technique compared with the CE VNTR analysis. The AE VNTR can therefore be regarded as a viable alternative in moderately equipped laboratories that cannot afford the expensive equipment required for CE VNTR analysis and data obtained by AE VNTR analysis can be shared between laboratories which use the CE VNTR method. (c) 2010 Elsevier B.V. All rights reserved.

  13. Studies of transferin polymorphism in Swedish cattle using agarose gel electrophoresis

    International Nuclear Information System (INIS)

    Liberg, P.; Carlstroem, G.

    1976-01-01

    The polymorphic transferrin picture in the sera from 894 Swedish cattle was investigated with an agarose gel electrophoresis technique. The serum transferrin bands in the electrophoresis pattern were first identified by labelling with 59 Fe. Six existing phenotypes based on the alleles Tf(supA), Tf(supD) and Tf(supE) could be detected. The frequencies of transferrin types and transferrin alleles are presented, and it is concluded that there are great differences in the frequencis between the Swedish Red and White and the Swedish Friesian. (author)

  14. Ionization techniques in capillary electrophoresis-mass spectrometry: principles, design, and application.

    Science.gov (United States)

    Hommerson, Paul; Khan, Amjad M; de Jong, Gerhardus J; Somsen, Govert W

    2011-01-01

    A major step forward in the development and application of capillary electrophoresis (CE) was its coupling to ESI-MS, first reported in 1987. More than two decades later, ESI has remained the principal ionization technique in CE-MS, but a number of other ionization techniques have also been implemented. In this review the state-of-the-art in the employment of soft ionization techniques for CE-MS is presented. First the fundamentals and general challenges of hyphenating conventional CE and microchip electrophoresis with MS are outlined. After elaborating on the characteristics and role of ESI, emphasis is put on alternative ionization techniques including sonic spray ionization (SSI), thermospray ionization (TSI), atmospheric pressure chemical ionization (APCI), atmospheric pressure photoionization (APPI), matrix-assisted laser desorption ionization (MALDI) and continuous-flow fast atom bombardment (CF-FAB). The principle of each ionization technique is outlined and the experimental set-ups of the CE-MS couplings are described. The strengths and limitations of each ionization technique with respect to CE-MS are discussed and the applicability of the various systems is illustrated by a number of typical examples. Copyright © 2011 Wiley Periodicals, Inc.

  15. Reference interval determination of hemoglobin fractions in umbilical cord and placental blood by capillary electrophoresis.

    Science.gov (United States)

    Bó, Suzane Dal; de Oliveira Lemos, Fabiane Kreutz; Pedrazzani, Fabiane Spagnol; Cagliari, Cláudia Rosa; Scotti, Luciana

    2016-04-01

    Umbilical cord and placental blood (UCPB) is a rich source of hematopoietic stem cells widely used to treat diseases that did not have effective treatments until recently. Umbilical cord and placental blood banks (UCPBBs) are needed to be created to store UCPB. UCPB is collected immediately after birth, processed, and frozen until infusion. Detection of abnormal hemoglobins is one of UCPB screening tests available. The objective of the present study was to determine the reference interval for HbA, HbF, and HbA2 in UCPB using capillary electrophoresis. Methods: Observational retrospective study of UCPB samples undergoing hemoglobin electrophoresis was performed between April 2012 and May 2013. We analyzed 273 UCPB samples. All cords met the criteria of BrasilCORD. We found 19.9% (10.5–36.7%) for HbA, 80.1% (62.7–89.4%) for HbF, and 0.1% (0.0–0.6%) for HbA2. Data were expressed as median (P2.5–P97.5). Establishing specific reference intervals is the best option for most tests because such ranges reflect the status of the population in which the tests will be applied. The use of appropriate reference intervals ensures that clinical labs provide reliable information, thus enabling clinicians to correctly interpret results and choose the best approach for the target population.

  16. 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.

  17. Determination of the R-enantiomer of valsartan in pharmaceutical formulation by capillary electrophoresis.

    Science.gov (United States)

    Lee, Kyung Ran; Nguyen, NgocVan Thi; Lee, Yong Jae; Choi, Seungho; Kang, Jong Seong; Mar, Woongchon; Kim, Kyeong Ho

    2015-01-01

    Capillary zone electrophoresis was successfully applied to the enantiomeric purity determination of valsartan using acetyl-β-cyclodextrin (A-β-CD) as a chiral selector. Separations were carried out in a 50 µm, 64/56 cm fused-silica capillary. The optimized conditions included 25 mM phosphate buffer, pH 8.0, containing 10 mM A-β-CD as background electrolyte, an applied voltage of +30 kV and a temperature of 30 °C. Ibuprofen was used as an internal standard. The assay was validated for the R-enantiomer of valsartan in the range of 0.05-3.0%. The limit of detection was 0.01%, the limit of quantitation was 0.05%, relative to a concentration of valsartan of 1 mg/ml. Intra-day precision varied between 2.57 and 5.60%. Relative standard deviations of inter-day precision ranged between 4.46 and 6.76% for peak area ratio. The percentage recovery of the R-enantiomer of valsartan ranged between 97.0 and 99.6% in valsartan product. The assay was applied to the determination of the chiral purity of valsartan tablets and R-enantiomer of valsartan was found as an impurity.

  18. Evaluation of Global Genomic DNA Methylation in Human Whole Blood by Capillary Electrophoresis UV Detection

    Directory of Open Access Journals (Sweden)

    Angelo Zinellu

    2017-01-01

    Full Text Available Alterations in global DNA methylation are implicated in various pathophysiological processes. The development of simple and quick, yet robust, methods to assess DNA methylation is required to facilitate its measurement and interpretation in clinical practice. We describe a highly sensitive and reproducible capillary electrophoresis method with UV detection for the separation and detection of cytosine and methylcytosine, after formic acid hydrolysis of DNA extracted from human whole blood. Hydrolysed samples were dried and resuspended with water and directly injected into the capillary without sample derivatization procedures. The use of a run buffer containing 50 mmol/L BIS-TRIS propane (BTP phosphate buffer at pH 3.25 and 60 mmol/L sodium acetate buffer at pH 3.60 (4 : 1, v/v allowed full analyte identification within 11 min. Precision tests indicated an elevated reproducibility with an interassay CV of 1.98% when starting from 2 μg of the extracted DNA. The method was successfully tested by measuring the DNA methylation degree both in healthy volunteers and in reference calf thymus DNA.

  19. On gel electrophoresis of dielectric charged particles with hydrophobic surface: A combined theoretical and numerical study.

    Science.gov (United States)

    Majee, Partha Sarathi; Bhattacharyya, Somnath; Gopmandal, Partha Pratim; Ohshima, Hiroyuki

    2018-03-01

    A theoretical study on the gel electrophoresis of a charged particle incorporating the effects of dielectric polarization and surface hydrophobicity at the particle-liquid interface is made. A simplified model based on the weak applied field and low charge density assumption is also presented and compared with the full numerical model for a nonpolarizable particle to elucidate the nonlinear effects such as double layer polarization and relaxation as well as surface conduction. The main motivation of this study is to analyze the electrophoresis of the surface functionalized nanoparticle with tunable hydrophobicity or charged fluid drop in gel medium by considering the electrokinetic effects and hydrodynamic interactions between the particle and the gel medium. An effective medium approach, in which the transport in the electrolyte-saturated hydrogel medium is governed by the Brinkman equation, is adopted in the present analysis. The governing electrokinetic equations based on the conservation principles are solved numerically. The Navier-slip boundary condition along with the continuity condition of dielectric displacement are imposed on the surface of the hydrophobic polarizable particle. The impact of the slip length on the electrophoresis is profound for a thinner Debye layer, however, surface conduction effect also becomes significant for a hydrophobic particle. Impact of hydrophobicity and relaxation effects are higher for a larger particle. Dielectric polarization creates a reduction in its electrophoretic propulsion and has negligible impact at the thinner Debye length as well as lower gel screening length. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Speciation of protein-bound trace elements by gel electrophoresis and atomic spectrometry.

    Science.gov (United States)

    Ma, Renli; McLeod, Cameron W; Tomlinson, Kerry; Poole, Robert K

    2004-08-01

    The metabolism of trace elements, in particular their binding to proteins in biological systems is of great importance in biochemical, toxicological, and pharmacological studies. As a result there has been a sustained interest over the last two decades in the speciation of protein-bound metals. Various analytical approaches have been employed, combining efficient separation of metalloproteins by liquid chromatography or electrophoresis with high-sensitivity elemental detection. Slab-gel electrophoresis (GE) is a key platform for high-resolution protein separation, and has been combined with autoradiography and various atomic spectrometric techniques for in-gel determination of protein-bound metals. Recently, the combination of GE with state-of-the-art inductively coupled plasma-mass spectrometry (ICP-MS), particularly when linked to laser ablation (LA) for direct gel interrogation, has opened up new opportunities for rapid characterization of metalloproteins. The use of GE and atomic spectrometry for the speciation of protein-bound trace elements is reviewed in this paper. Technical requirements for gel electrophoresis/atomic spectrometric measurement are considered in terms of method compatibilities, detection capability and potential usefulness. The literature is also surveyed to illustrate current status and future trends. Copyright 2004 Wiley-VCH Verlag GmbH and Co.

  1. Peak capacity and peak capacity per unit time in capillary and microchip zone electrophoresis.

    Science.gov (United States)

    Foley, Joe P; Blackney, Donna M; Ennis, Erin J

    2017-11-10

    The origins of the peak capacity concept are described and the important contributions to the development of that concept in chromatography and electrophoresis are reviewed. Whereas numerous quantitative expressions have been reported for one- and two-dimensional separations, most are focused on chromatographic separations and few, if any, quantitative unbiased expressions have been developed for capillary or microchip zone electrophoresis. Making the common assumption that longitudinal diffusion is the predominant source of zone broadening in capillary electrophoresis, analytical expressions for the peak capacity are derived, first in terms of migration time, diffusion coefficient, migration distance, and desired resolution, and then in terms of the remaining underlying fundamental parameters (electric field, electroosmotic and electrophoretic mobilities) that determine the migration time. The latter expressions clearly illustrate the direct square root dependence of peak capacity on electric field and migration distance and the inverse square root dependence on solute diffusion coefficient. Conditions that result in a high peak capacity will result in a low peak capacity per unit time and vice-versa. For a given symmetrical range of relative electrophoretic mobilities for co- and counter-electroosmotic species (cations and anions), the peak capacity increases with the square root of the electric field even as the temporal window narrows considerably, resulting in a significant reduction in analysis time. Over a broad relative electrophoretic mobility interval [-0.9, 0.9], an approximately two-fold greater amount of peak capacity can be generated for counter-electroosmotic species although it takes about five-fold longer to do so, consistent with the well-known bias in migration time and resolving power for co- and counter-electroosmotic species. The optimum lower bound of the relative electrophoretic mobility interval [μ r,Z , μ r,A ] that provides the maximum

  2. Open microfluidic gel electrophoresis: Rapid and low cost separation and analysis of DNA at the nanoliter scale.

    Science.gov (United States)

    Gutzweiler, Ludwig; Gleichmann, Tobias; Tanguy, Laurent; Koltay, Peter; Zengerle, Roland; Riegger, Lutz

    2017-07-01

    Gel electrophoresis is one of the most applied and standardized tools for separation and analysis of macromolecules and their fragments in academic research and in industry. In this work we present a novel approach for conducting on-demand electrophoretic separations of DNA molecules in open microfluidic (OM) systems on planar polymer substrates. The approach combines advantages of slab gel, capillary- and chip-based methods offering low consumable costs (<0.1$) circumventing cost-intensive microfluidic chip fabrication, short process times (5 min per analysis) and high sensitivity (4 ng/μL dsDNA) combined with reasonable resolution (17 bases). The open microfluidic separation system comprises two opposing reservoirs of 2-4 μL in volume, a semi-contact written gel line acting as separation channel interconnecting the reservoirs and sample injected into the line via non-contact droplet dispensing and thus enabling the precise control of the injection plug and sample concentration. Evaporation is prevented by covering aqueous structures with PCR-grade mineral oil while maintaining surface temperature at 15°C. The liquid gel line exhibits a semi-circular cross section of adaptable width (∼200-600 μm) and height (∼30-80 μm) as well as a typical length of 15-55 mm. Layout of such liquid structures is adaptable on-demand not requiring time consuming and repetitive fabrication steps. The approach was successfully demonstrated by the separation of a standard label-free DNA ladder (100-1000 bp) at 100 V/cm via in-line staining and laser induced fluorescent end-point detection using an automated prototype. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Extracting information from two-dimensional electrophoresis gels by partial least squares regression

    DEFF Research Database (Denmark)

    Jessen, Flemming; Lametsch, R.; Bendixen, E.

    2002-01-01

    of all proteins/spots in the gels. In the present study it is demonstrated how information can be extracted by multivariate data analysis. The strategy is based on partial least squares regression followed by variable selection to find proteins that individually or in combination with other proteins vary......Two-dimensional gel electrophoresis (2-DE) produces large amounts of data and extraction of relevant information from these data demands a cautious and time consuming process of spot pattern matching between gels. The classical approach of data analysis is to detect protein markers that appear...... or disappear depending on the experimental conditions. Such biomarkers are found by comparing the relative volumes of individual spots in the individual gels. Multivariate statistical analysis and modelling of 2-DE data for comparison and classification is an alternative approach utilising the combination...

  4. Increase in local protein concentration by field-inversion gel electrophoresis

    Directory of Open Access Journals (Sweden)

    Paulus Aran

    2007-09-01

    Full Text Available Abstract Background Proteins that migrate through cross-linked polyacrylamide gels (PAGs under the influence of a constant electric field experience negative factors, such as diffusion and non-specific trapping in the gel matrix. These negative factors reduce protein concentrations within a defined gel volume with increasing migration distance and, therefore, decrease protein separation efficiency. Enhancement of protein separation efficiency was investigated by implementing pulsed field-inversion gel electrophoresis (FIGE. Results Separation of model protein species and large protein complexes was compared between FIGE and constant field electrophoresis (CFE in different percentages of PAGs. Band intensities of proteins in FIGE with appropriate ratios of forward and backward pulse times were superior to CFE despite longer running times. These results revealed an increase in band intensity per defined gel volume. A biphasic protein relative mobility shift was observed in percentages of PAGs up to 14%. However, the effect of FIGE on protein separation was stochastic at higher PAG percentage. Rat liver lysates subjected to FIGE in the second-dimension separation of two-dimensional polyarcylamide gel electrophoresis (2D PAGE showed a 20% increase in the number of discernible spots compared with CFE. Nine common spots from both FIGE and CFE were selected for peptide sequencing by mass spectrometry (MS, which revealed higher final ion scores of all nine protein spots from FIGE. Native protein complexes ranging from 800 kDa to larger than 2000 kDa became apparent using FIGE compared with CFE. Conclusion The present investigation suggests that FIGE under appropriate conditions improves protein separation efficiency during PAGE as a result of increased local protein concentration. FIGE can be implemented with minimal additional instrumentation in any laboratory setting. Despite the tradeoff of longer running times, FIGE can be a powerful protein

  5. Increase in local protein concentration by field-inversion gel electrophoresis.

    Science.gov (United States)

    Tsai, Henghang; Low, Teck Yew; Freeby, Steve; Paulus, Aran; Ramnarayanan, Kalpana; Cheng, Chung-Pui Paul; Leung, Hon-Chiu Eastwood

    2007-09-26

    Proteins that migrate through cross-linked polyacrylamide gels (PAGs) under the influence of a constant electric field experience negative factors, such as diffusion and non-specific trapping in the gel matrix. These negative factors reduce protein concentrations within a defined gel volume with increasing migration distance and, therefore, decrease protein separation efficiency. Enhancement of protein separation efficiency was investigated by implementing pulsed field-inversion gel electrophoresis (FIGE). Separation of model protein species and large protein complexes was compared between FIGE and constant field electrophoresis (CFE) in different percentages of PAGs. Band intensities of proteins in FIGE with appropriate ratios of forward and backward pulse times were superior to CFE despite longer running times. These results revealed an increase in band intensity per defined gel volume. A biphasic protein relative mobility shift was observed in percentages of PAGs up to 14%. However, the effect of FIGE on protein separation was stochastic at higher PAG percentage. Rat liver lysates subjected to FIGE in the second-dimension separation of two-dimensional polyarcylamide gel electrophoresis (2D PAGE) showed a 20% increase in the number of discernible spots compared with CFE. Nine common spots from both FIGE and CFE were selected for peptide sequencing by mass spectrometry (MS), which revealed higher final ion scores of all nine protein spots from FIGE. Native protein complexes ranging from 800 kDa to larger than 2000 kDa became apparent using FIGE compared with CFE. The present investigation suggests that FIGE under appropriate conditions improves protein separation efficiency during PAGE as a result of increased local protein concentration. FIGE can be implemented with minimal additional instrumentation in any laboratory setting. Despite the tradeoff of longer running times, FIGE can be a powerful protein separation tool.

  6. Genetic heterogeneity of Campylobacter concisus determined by pulsed field gel electrophoresis-based macrorestriction profiling

    DEFF Research Database (Denmark)

    Matsheka, M.I.; Elisha, B.G.; Lastovica, A.L.

    2002-01-01

    1 for pulsed field gel electrophoresis-based genotyping. Subsequently, 53 strains of C concisus, principally from cases of diarrhoea in children, were examined. Fifty-one distinct patterns were obtained, indicating the high discriminatory potential of the method. Patterns comprised between one...... comprised of several genomospecies. The pulsed field gel electrophoresis typing method described here has considerable potential for molecular epidemiological studies of C concisus and may be a useful adjunctive method for helping to resolve key taxonomic issues for this species....... and 14 restriction fragments, with type and reference strains of two well-defined genomospecies of oral and faecal origin containing six and 12 fragments respectively. Our results show that C concisus is genetically diverse and suggest the species as currently defined to be a taxonomic continuum...

  7. Salmonella enterica pulsed-field gel electrophoresis clusters, Minnesota, USA, 2001-2007.

    Science.gov (United States)

    Rounds, Joshua M; Hedberg, Craig W; Meyer, Stephanie; Boxrud, David J; Smith, Kirk E

    2010-11-01

    We determined characteristics of Salmonella enterica pulsed-field gel electrophoresis clusters that predict their being solved (i.e., that result in identification of a confirmed outbreak). Clusters were investigated by the Minnesota Department of Health by using a dynamic iterative model. During 2001-2007, a total of 43 (12.5%) of 344 clusters were solved. Clusters of ≥4 isolates were more likely to be solved than clusters of 2 isolates. Clusters in which the first 3 case isolates were received at the Minnesota Department of Health within 7 days were more likely to be solved than were clusters in which the first 3 case isolates were received over a period >14 days. If resources do not permit investigation of all S. enterica pulsed-field gel electrophoresis clusters, investigation of clusters of ≥4 cases and clusters in which the first 3 case isolates were received at a public health laboratory within 7 days may improve outbreak investigations.

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

    International Nuclear Information System (INIS)

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

    1985-01-01

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

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

    OpenAIRE

    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...

  10. Pulsed-field Gel Electrophoresis for Salmonella Infection Surveillance, Texas, USA, 2007

    Centers for Disease Control (CDC) Podcasts

    2010-06-14

    This podcast describes monitoring of the use of pulsed-field gel electrophoresis for Salmonella surveillance in Houston, Texas. CDC microbiologist Peter Gerner-Smidt discusses the importance of the PulseNet national database in surveillance of food-borne infections.  Created: 6/14/2010 by National Center for Emerging and Zoonotic Infectious Diseases (NCEZID).   Date Released: 6/14/2010.

  11. Trapping and breaking of in vivo nicked DNA during pulsed-field gel electrophoresis

    Science.gov (United States)

    Khan, Sharik R.; Kuzminov, Andrei

    2013-01-01

    Pulsed field gel electrophoresis (PFGE) offers a high-resolution approach to quantify chromosomal fragmentation in bacteria, measured as percent of chromosomal DNA entering the gel. The degree of separation in PFG depends upon the size of DNA, as well as various conditions of electrophoresis, such as electric field strength (FS), time of electrophoresis, switch time and buffer composition. Here we describe a new parameter, the structural integrity of the sample DNA itself, that influences its migration through PFGs. We show that sub-chromosomal fragments containing both spontaneous and DNA damage-induced nicks are prone to breakage during PFGE. Such breakage at single strand interruptions results in artefactual decrease in molecular weight of linear DNA making accurate determination of the number of double strand breaks difficult. While breakage of nicked sub-chromosomal fragments is FS-independent, some high molecular weight sub-chromosomal fragments are also trapped within wells under the standard PFGE conditions. This trapping can be minimized by lowering the field strength and increasing the time of electrophoresis. We discuss how breakage of nicked DNA may be mechanistically linked to trapping. Our results suggest how to optimize conditions for PFGE when quantifying chromosomal fragmentation induced by DNA damage. PMID:23770235

  12. Alkaline gel electrophoresis assay to detect DNA strand breaks and repair mechanisms in Escherichia coli

    Energy Technology Data Exchange (ETDEWEB)

    Mattos, Jose Carlos Pelielo de; Motta, Ellen Serri da; Oliveira, Marcia Betania Nunes de; Dantas, Flavio Jose da Silva; Araujo, Adriano Caldeira de [Universidade do Estado do Rio de Janeiro (UERJ), RJ (Brazil). Dept. de Biofisica e Biometria. Lab. de Radio e Fotobiologia]. E-mail: jcmattos@uerj.br

    2008-12-15

    Reactive oxygen species (ROS) can induce lesions in different cellular targets, including DNA. Stannous chloride (SnCl{sub 2}) is a ROS generator, leading to lethality in Escherichia coli (E. coli), with the base excision repair (BER) mechanism playing a role in this process. Many techniques have been developed to detect genotoxicity, as comet assay, in eukaryotic cells, and plasmid DNA agarose gel electrophoresis. In this study, an adaptation of the alkaline gel electrophoresis method was carried out to ascertain the induction of strand breaks by SnCl{sub 2} in bacterial DNA, from E. coli BER mutants, and its repair pathway. Results obtained show that SnCl{sub 2} was able to induce DNA strand breaks in all strains tested. Moreover, endonuclease IV and exonuclease III play a role in DNA repair. On the whole, data has shown that the alkaline gel electrophoresis assay could be used both for studying DNA strand breaks induction and for associated repair mechanisms. (author)

  13. Application of capillary electrophoresis to the simultaneous determination and stability study of four extensively used penicillin derivatives

    Directory of Open Access Journals (Sweden)

    Brigitta Simon

    2014-09-01

    Full Text Available The applicability of capillary electrophoresis for the analysis of four extensively used penicillin derivatives (benzylpenicillin, ampicillin, amoxicillin, oxacilllin has been studied. Because of structural similarities, the electrophoretic behavior of these derivatives is very similar; consequently an efficient separation using the conventional capillary zone electrophoresis is hard to be achieved. Their simultaneous separation was solved by using micellar electrokinetic capillary chromatography, the separation being based on the differential partition of the analytes between the micellar and aqueous phase. Using a buffer solution containing 25 mM sodium tetraborate and 100 mM sodium dodecyl sulfate as surfactant, at a pH of 9.3, applying a voltage of + 25 kV at a temperature of 25 °C, we achieved the simultaneous separation of the studied penicillin derivatives in less then 5 minutes. The separation conditions were optimized and the analytical performance of the method was evaluated in terms of precision, linearity, limit of detection, and quantification. Also, a simple capillary zone electrophoresis method was applied to study the stability of the studied penicillin derivatives in water at different temperatures, using ciprofloxacin hydrochloride as internal standard. It was observed that the extent of the hydrolysis of penicillins in water is highly dependent on the time and also temperature.

  14. Automation and integration of polymerase chain reaction with capillary electrophoresis for high throughput genotyping and disease diagnosis

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, N.

    1999-02-12

    Genotyping is to detect specific loci in the human genome. These loci provide important information for forensic testing, construction of genetic linkage maps, gene related disease diagnosis and pharmacogenetic research. Genotyping is becoming more and more popular after these loci can be easily amplified by polymerase chain reaction (PCR). Capillary electrophoresis has its unique advantages for DNA analysis due to its fast heat dissipation and ease of automation. Four projects are described in which genotyping is performed by capillary electrophoresis emphasizing different aspects. First, the author demonstrates a principle to determine the genotype based on capillary electrophoresis system. VNTR polymorphism in the human D1S80 locus was studied. Second, the separation of four short tandem repeat (STR) loci vWF, THO1, TPOX and CSF1PO (CTTv) by using poly(ethylene oxide) (PEO) was studied in achieving high resolution and preventing rehybridization of the DNA fragments. Separation under denaturing, non-denaturing conditions and at elevated temperature was discussed. Third, a 250 {micro}m i.d., 365 {micro}m o.d. fused silica capillary was used as the microreactor for PCR. Fourth, direct PCR from blood was studied to simplify the sample preparation for genotyping to minimum.

  15. A critical overview of non-aqueous capillary electrophoresis. Part II: separation efficiency and analysis time.

    Science.gov (United States)

    Kenndler, Ernst

    2014-03-28

    A survey of the literature on non-aqueous capillary zone electrophoresis leaves one with the impression of a prevailing notion that non-aqueous conditions are principally more favorable than conventional aqueous media. Specifically, the application of organic solvents in capillary zone electrophoresis (CZE) is believed to provide the general advantages of superior separation efficiency, higher applicable electric field strength, and shorter analysis time. These advantages, however, are often claimed without providing any experimental evidence, or based on rather uncritical comparisons of limited sets of arbitrarily selected separation results. Therefore, the performance characteristics of non-aqueous vs. aqueous CZE certainly deserve closer scrutiny. The primary intention of Part II of this review is to give a critical survey of the literature on non-aqueous capillary electrophoresis (NACE) that has emerged over the last five years. Emphasis is mainly placed on those studies that are concerned with the aspects of plate height, plate number, and the crucial mechanisms contributing to zone broadening, both in organic and aqueous conditions. To facilitate a deeper understanding, this treatment covers also the theoretical fundamentals of peak dispersion phenomena arising from wall adsorption; concentration overload (electromigration dispersion); longitudinal diffusion; and thermal gradients. Theoretically achievable plate numbers are discussed, both under limiting (at zero ionic strength) and application-relevant conditions (at finite ionic strength). In addition, the impact of the superimposed electroosmotic flow contributions to overall CZE performance is addressed, both for aqueous and non-aqueous media. It was concluded that for peak dispersion due to wall adsorption and due to concentration overload (electromigration dispersion, leading to peak triangulation) no general conjunction with the solvent can be deduced. This is in contrast to longitudinal diffusion: the

  16. [Establishment of two-dimensional differential gel electrophoresis using cerebrospinal fluid from neurocysticercosis patients].

    Science.gov (United States)

    Li, Jing-Yi; Tian, Xiao-Jun; Huang, Yong; Yang, Yan-Jun; Ma, Qiao-Rong; Xue, Yan-Ping

    2008-06-30

    To establish the method of two-dimensional differential gel electrophoresis and obtain high resolution 2D images from cerebrospinal fluid (CSF) of patients with neurocysticercosis. CSF samples were collected from four patients diagnosed as neurocysticercosis clinically and by ELISA, computed tomography (CT) or magnetic resonance imaging (MRI), and from four healthy subjects without neurological disorders. The CSF samples were precipitated with cold acetone, then pooled by equal amount as patients and controls. The internal standard comprised equal amounts of proteins extracted from both groups. Internal standard, and proteins from the two groups were labeled prior to electrophoresis with spectrally resolvable fluorescent dyes, cyanein dye2 (Cy2), Cy3 and Cy5. Sodium dodecylsulfonate polyacrylamide gel chromatography (SDS-PAGE) and two-dimensional differential in-gel electrophoresis (2-D DIGE) of labeled samples were then run. The differential expressed proteins showed in the images of SDS-PAGE and 2-D DIGE gels scanned with 488 nm, 532 nm and 633 nm wavelength laser were analyzed by ImageQuant and DeCyde 5.0 respectively. Spot detection and quantification was performed for the differential in-gel analysis (DIA) module of DeCyder. Biological variation analysis (BVA) module of DeCyder was matched gel 1 and gel 2 images to provide data on differential protein expression levels between the two groups. The ImageQuant result displayed that the CSF protein was compatible with the dye, and the difference of protein amount was revealed by the difference of fluorescence intensity. DIA indicated that there were 896 and 894 protein dots on gel 1 and gel 2 respectively, and 90% of them were matched each other. BVA showed that there were 55 protein spots with different expressional level between neurocysticercosis and control groups. Protein spots with two-fold increase or decrease were 47 and 8 respectively in neurocysticercosis patients compared with healthy controls. The

  17. 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.

  18. 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.

  19. Determination of 2-methylimidazole and 4-methylimidazole in caramel colors by capillary electrophoresis.

    Science.gov (United States)

    Petruci, João Flávio da Silveira; Pereira, Elisabete Alves; Cardoso, Arnaldo Alves

    2013-03-06

    The use of chemical preservative compounds is common in the food products industry. Caramel color is the most usual additive used in beverages, desserts, and breads worldwide. During its fabrication process, 2- and 4-methylimidazole (MeI), highly carcinogenic compounds, are generated. In these cases, the development of reliable analytical methods for the monitoring of undesirable compounds is necessary. The primary procedure for the analysis of 2- and 4-MeI is using LC- or GC-MS techniques. These procedures are time-consuming and require large amounts of organic solvents and several pretreatment steps. This prevents the routine use of this procedure. This paper describes a rapid, efficient, and simple method using capillary electrophoresis (CE) for the separation and determination of 2- and 4-MeI in caramel colors. The analyses were performed using a 75 μm i.d. uncoated fused-silica capillary with an effective length of 40 cm and a running electrolyte consisting of 160 mmol L(-1) phosphate plus 30% acetonitrile. The pH was adjusted to 2.5 with triethylamine. The analytes were separated within 6 min at a voltage of 20 kV. Method validation revealed good repeatability of both migration time (<0.8% RSD) and peak area (<2% RSD). Analytical curves for 2- and 4-MeI were linear in the 0.4-40 mg L(-1) concentration interval. Detection limits were 0.16 mg L(-1) for 4-MeI and 0.22 mg L(-1) for 2-MeI. The extraction recoveries were satisfactory. The developed method showed many advantages when compared to the previously used method.

  20. Determination of lipoic acid in human urine by capillary zone electrophoresis.

    Science.gov (United States)

    Kubalczyk, Paweł; Głowacki, Rafał

    2017-07-01

    Fast, simple, and accurate CE method enabling determination of lipoic acid (LA) in human urine has been developed and validated. LA is a disulfide-containing natural compound absorbed from the organism's diet. Due to powerful antioxidant activity, LA has been used for prevention and treatment of various diseases and disorders, e.g. cardiovascular diseases, neurodegenerative disorders, and cancer. The proposed analytical procedure consists of liquid-liquid sample extraction, reduction of LA with tris(2-carboxyethyl)phosphine, derivatization with 1-benzyl-2-chloropyridinium bromide (BCPB) followed by field amplified sample injection stacking, capillary zone electrophoresis separation, and ultraviolet-absorbance detection of LA-BCPB derivative at 322 nm. Effective baseline electrophoretic separation was achieved within 6 min under the separation voltage of 20 kV (∼80 μA) using a standard fused-silica capillary (effective length 51.5 cm, 75 μm id) and BGE consisted of 0.05 mol/L borate buffer adjusted to pH 9. The experimentally determined limit of detection for LA in urine was 1.2 μmol/L. The calibration curve obtained for LA in urine showed linearity in the range 2.5-80 μmol/L, with R 2 0.9998. The relative standard deviation of the points of the calibration curve was lower than 10%. The analytical procedure was successfully applied to analysis of real urine samples from seven healthy volunteers who received single 100 mg dose of LA. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Analytical potential of mid-infrared detection in capillary electrophoresis and liquid chromatography: A review

    International Nuclear Information System (INIS)

    Kuligowski, Julia; Quintas, Guillermo; Guardia, Miguel de la; Lendl, Bernhard

    2010-01-01

    Literature published in the last decade concerning the use of mid-infrared spectrometry as a detection system in separation techniques employing a liquid mobile phase is reviewed. In addition to the continued use of isocratic liquid chromatographic (LC) techniques, advances in chemometric data evaluation techniques now allow the use of gradient techniques on a routine basis, thus significantly broadening the range of possible applications of LC-IR. The general trend towards miniaturized separation systems was also followed for mid-IR detection where two key developments are of special importance. Firstly, concerning on-line detection the advent of micro-fabricated flow-cells with inner volumes of only a few nL for transmission as well as attenuated total reflection measurements enabled on-line mid-IR detection in capillary LC and opened the path for the first successful realization of on-line mid-IR detection in capillary zone electrophoresis as well as micellar electrokinetic chromatography. Secondly, concerning off-line detection the use of micro-flow through dispensers now enables to concentrate eluting analytes on dried spots sized a few tens of micrometers, thus matching the dimensions for sensitive detection by mid-IR microscopy. Finally in an attempt to increase detection sensitivity of on-line mid-IR detection, mid-IR quantum cascade lasers have been used. Applications cover the field of food analysis, environmental analysis and the characterization of explosives among others. Best detection sensitivities for on-line and off-line detection have been achieved in miniaturized systems and are in the order of 50 ng and 2 ng on column, respectively.

  2. Quasi-isoelectric buffers for protein analysis in a fast alternative to conventional capillary zone electrophoresis.

    Science.gov (United States)

    Antonioli, Paolo; Mendieta, Martha E; Sebastiano, Roberto; Citterio, Attilio; Peltre, Gabriel; Busnel, Jean-Marc; Descroix, Stephanie; Candiano, Giovanni; Righetti, Pier Giorgio

    2006-03-20

    Two different approaches are here reported for obtaining ultra-narrow pI cuts from 2-pH unit wide carrier ampholyte ranges, as commercially available, for use as quasi-isoelectric buffers in capillary electrophoresis separations of proteins. One of them uses multicompartment electrolyzers endowed with isoelectric membranes (Immobiline technology); the other employs the Rotofor equipment. Although the first approach results in more precise pI cuts, the latter technique is much faster, easier to handle and permits the immediate collection of 20 fractions in a single run. This results in ultra-narrow, ca. 0.1-pH unit intervals, uniformly spaced apart along the original wider gradient utilized for the fractionation. It is here shown that such quasi-isoelectric buffers, especially those in the pH 8-9 interval, have the unique property of coating the silica wall, thus preventing interaction of the proteins with the silica surface, that would otherwise totally disrupt the separation. On the contrary, such a shielding is not obtained in control, non isoelectric buffers (such as phosphate), that give very poor separations in uncoated capillaries. It is hypothesized that such a unique shielding effect is due to the oligo-amino backbone of the carrier ampholytes, typically composed (in the Vesterberg's synthetic approach) of 4-6 nitrogens spaced apart by ethylene moieties. Although such oligoprotic buffers should bear, in the isoelectric state, just one positive and one negative charge, they might be transiently ionized upon contact with the silanols, thus inducing a cooperative binding to the silica wall.

  3. Monitoring the enzymatic conversion of urea to ammonium by conventional or microchip capillary electrophoresis with contactless conductivity detection.

    Science.gov (United States)

    Schuchert-Shi, Aiping; Hauser, Peter C

    2008-05-15

    Capillary electrophoresis with contactless conductivity detection was used to directly quantify the ammonium produced in the enzymatic conversion of urea with urease. This allowed the characterization of the reaction without having to use more elaborate indirect optical methods for quantification. The maximum rate of reaction, V(max), was determined as 5.1 mmol x mL(-1) x min(-1), and the Michaelis-Menten constant, K(m), was determined as 16 mM. Furthermore, the method was successfully applied to the determination of urea in clinical samples of human blood by using a conventional capillary and a microchip device.

  4. A CCD-based system for the detection of DNA in electrophoresis gels by UV absorption

    International Nuclear Information System (INIS)

    Mahon, A.R.; MacDonald, J.H.; Mainwood, A.; Ott, R.J.

    1999-01-01

    A method and apparatus for the detection and quantification of large fragments of unlabelled nucleic acids in agarose gels is presented. The technique is based on ultraviolet (UV) absorption by nucleotides. A deuterium source illuminates individual sample lanes of an electrophoresis gel via an array of optical fibres. As DNA bands pass through the illuminated region of the gel the amount of UV light transmitted is reduced because of absorption by the DNA. During electrophoresis the regions of DNA are detected on-line using a UV-sensitive charge coupled device (CCD). As the absorption coefficient is proportional to the mass of DNA the technique is inherently quantitative. The mass of DNA in a region of the gel is approximately proportional to the integrated signal in the corresponding section of the CCD image. This system currently has a detection limit of less than 1.25 ng compared with 2-10 ng for the most popular conventional technique, ethidium bromide (EtBr) staining. In addition the DNA sample remains in its native state. The removal of the carcinogenic dye from the detection procedure greatly reduces associated biological hazards. (author)

  5. Determination of size distribution of small DNA fragments by polyacrylamide gel electrophoresis

    International Nuclear Information System (INIS)

    Lau How Mooi

    1998-01-01

    Size distribution determination of DNA fragments can be normally determined by the agarose gel electrophoresis, including the normal DNA banding pattern analysis. However this method is only good for large DNA, such as the DNA of the size of kilo base pairs to mega base pairs range. DNA of size less than kilo base pairs is difficult to be quantified by the agarose gel method. Polyacrylamide gel electrophoresis however can be used to measure the quantity of DNA fragments of size less than kilo base pairs in length, down to less than ten base pairs. This method is good for determining the quantity of the smaller size DNA, single stranded polymers or even some proteins, if the known standards are available. In this report detail description of the method of preparing the polyacrylamide gel, and the experimental set up is discussed. Possible uses of this method, and the comparison with the standard sizes of DNA is also shown. This method is used to determine the distribution of the amount of the fragmented DNA after the Calf-thymus DNA has been exposed to various types of radiation and of different doses. The standards were used to determine the sizes of the fragmented Calf-thymus DNA. The higher the dose the higher is the amount of the smaller size DNA measured

  6. A sol-gel-modified poly(methyl methacrylate) electrophoresis microchip with a hydrophilic channel wall.

    Science.gov (United States)

    Chen, Gang; Xu, Xuejiao; Lin, Yuehe; Wang, Joseph

    2007-01-01

    A sol-gel method was employed to fabricate a poly(methyl methacrylate) (PMMA) electrophoresis microchip that contains a hydrophilic channel wall. To fabricate such a device, tetraethoxysilane (TEOS) was injected into the PMMA channel and was allowed to diffuse into the surface layer for 24 h. After removing the excess TEOS, the channel was filled with an acidic solution for 3 h. Subsequently, the channel was flushed with water and was pretreated in an oven to obtain a sol-gel-modified PMMA microchip. The water contact angle for the sol-gel-modified PMMA was approximately 27.4 degrees compared with approximately 66.3 degrees for the pure PMMA. In addition, the electro-osmotic flow increased from 2.13x10(-4) cm2 V(-1) s(-1) for the native-PMMA channel to 4.86x10(-4) cm2 V(-1) s(-1) for the modified one. The analytical performance of the sol-gel-modified PMMA microchip was demonstrated for the electrophoretic separation of several purines, coupled with amperometric detection. The separation efficiency of uric acid increased to 74,882.3 m(-1) compared with 14,730.5 m(-1) for native-PMMA microchips. The result of this simple modification is a significant improvement in the performance of PMMA for microchip electrophoresis and microfluidic applications.

  7. Lesion measurement in non-radioactive DNA by quantitative gel electrophoresis

    International Nuclear Information System (INIS)

    Sutherland, J.C.; Chen, Chun Zhang; Emrick, A.; Hacham, H; Monteleone, D.; Ribeiro, E.; Trunk, J.; Sutherland, B.M.

    1989-01-01

    The gel electrophoresis method developed during the past ten years in our laboratories makes possible the quantitation of UV induced pyrimidine dimers, gamma ray induced single- and double-strand breaks and many other types of lesions in nanogram quantities of DNA. The DNA does not have to be labeled with radionuclides or of a particular conformation, thus facilitating the use of the method in measuring damage levels and repair rates in the DNA of intact organisms -- including man. The gel method can quantitate any lesion in DNA that either is, or can be converted to a single- or double-strand break. The formation of a strand break produces two shorter DNA molecules for each molecule that existed before the treatment that produced the break. Determining the number of breaks, and hence the number of lesions, becomes a matter of comparing the average lengths of molecules in samples differing only in lesion-induced breaks. This requires that we determine the distribution of mass of DNA on a gel as a function of its distance of migration and also the dispersion function of its distance of migration and also the dispersion function (the relationship between molecular length and distance of migration) in the gel electrophoresis system. 40 refs., 5 figs

  8. Sol-gel open tubular ODS columns with reversed electroosmotic flow for capillary electrochromatography.

    Science.gov (United States)

    Hayes, J D; Malik, A

    2001-03-01

    Sol-gel chemistry was successfully used for the fabrication of open tubular columns with surface-bonded octadecylsilane (ODS) stationary-phase coating for capillary electrochromatography (OT-CEC). Following column preparations, a series of experiments were performed to investigate the performance of the sol-gel coated ODS columns in OT-CEC. The incorporation of N-octadecyldimethyl[3-(trimethoxysilyl)propyl]ammonium chloride as one of the sol-gel precursors played an important role in the electrochromatographic performance of the prepared columns. This chemical reagent possesses a chromatographically favorable, bonded ODS moiety, in conjunction with three methoxy groups allowing for sol-gel reactivity. In addition, a positively charged nitrogen atom is present in the molecular structure of this reagent and provides a positively charged capillary surface responsible for the reversed electroosmotic flow (EOF) in the columns during CEC operation. Comparative studies involving the EOF within such sol-gel ODS coated and uncoated capillaries were performed using acetonitrile and methanol as the organic modifiers in the mobile phase. The use of a deactivating reagent, phenyldimethylsilane, in the sol-gel solution was evaluated. Efficiency values of over 400,000 theoretical plates per meter were achieved in CEC on a 64 cm x 25 microm i.d. sol-gel ODS open tubular column. Test mixtures of polycyclic aromatic hydrocarbons, benzene derivatives, and aromatic aldehydes and ketones were used to evaluate the CEC performances of both nondeactivated and deactivated open tubular sol-gel columns. The effects of mobile-phase organic modifier contents and pH on EOF in such columns were evaluated. The prepared sol-gel ODS columns are characterized by switchable electroosmotic flow. A pH value of approximately 8.5 was found correspond to the isoelectric point for the prepared sol-gel ODS coatings.

  9. Multivariate data analysis of two-dimensional gel electrophoresis protein patterns from few samples

    DEFF Research Database (Denmark)

    Jensen, Kristina Nedenskov; Jessen, Flemming; Jørgensen, Bo

    2008-01-01

    One application of 2D gel electrophoresis is to reveal differences in protein pattern between two or more groups of individuals, attributable to their group membership. Multivariate data analytical methods are useful in pinpointing the spots relevant for discrimination by focusing not only...... on single spot differences, but on the covariance structure between proteins. However, their outcome is dependent on data scaling, and they may fail in producing valid multivariate models due to the much higher number of "irrelevant" spots present in the gels. The case where only few gels are available...... and where the aim is to find as many as possible of the group-dependent proteins seems particularly difficult to handle. The present paper investigates such a case regarding the effect of scaling and of prefiltering by univariate nonparametric statistics on the selection of spots. Besides, a modified...

  10. Two-dimensional gel electrophoresis of selenized yeast and autoradiography of 75Se-containing proteins

    International Nuclear Information System (INIS)

    Chery, C.C.; Dumont, E.; Cornelis, R.; Moens, L.

    2001-01-01

    Two-dimensional high-resolution gel electrophoresis (2DE) has been applied to the fractionation of 75 Se-containing proteins in yeast, grown in 75 Se-containing medium, and autoradiography was used for detection of the 75 Se-containing proteins. Gel filtration and ultrafiltration were used to check whether the selenium side-chains were stable in the presence of the chemicals used for lysis and 2DE. The mass distribution of the selenium-containing proteins was estimated by use of gel filtration and the results were compared with the distribution obtained by 2DE. A 2DE map of selenium-containing proteins in yeast is presented, and compared with a total protein map of yeast. (orig.)

  11. [Analysis of tartrazine aluminum lake and sunset yellow aluminum lake in foods by capillary zone electrophoresis].

    Science.gov (United States)

    Zhang, Yiding; Chang, Cuilan; Guo, Qilei; Cao, Hong; Bai, Yu; Liu, Huwei

    2014-04-01

    A novel analytical method for tartrazine aluminum lake and sunset yellow aluminum lake using capillary zone electrophoresis (CZE) was studied. The pigments contained in the color lakes were successfully separated from the aluminum matrix in the pre-treatment process, which included the following steps: dissolve the color lakes in 0.1 mol/L H2SO4, adjust the pH of the solution to 5.0, then mix it with the solution of EDTA x 2Na and heat it in a water bath, then use polyamide powder as the stationary phase of solid phase extraction to separate the pigments from the solution, and finally elute the pigments with 0.1 mol/L NaOH. The CZE conditions systematically optimized for tartrazine aluminum lake were: 48.50 cm of a fused silica capillary with 40.00 cm effective length and 50 microm i. d., the temperature controlled at 20.0 degrees C, 29.0 kV applied, HPO4(2-)-PO4(3-) (0.015 mol/L, pH 11.45) solution as running buffer, detection at 263 nm. The conditions for sunset yellow aluminum lake were: the same capillary and temperature, 25.0 kV applied, HPO4(2-)-PO4(3-) (0.025 mol/L, pH 11.45) solution as running buffer, detection at 240 nm. The limits of detection were 0.26 mg/L and 0.27 mg/L, and the linear ranges were 0.53-1.3 x 10(2) mg/L and 0.54-1.4 x 10(2) mg/L for tartrazine aluminum lake and sunset yellow aluminum lake, respectively. The RSDs were 4.3% and 5.7% (run to run, n = 6), 5.6% and 6.0% (day to day, n = 6) for tartrazine aluminum lake and sunset yellow aluminum lake, respectively. Further developments for this method could make it a routinely used method analyzing color lakes in foods.

  12. Smartphone Cortex Controlled Real-Time Image Processing and Reprocessing for Concentration Independent LED Induced Fluorescence Detection in Capillary Electrophoresis.

    Science.gov (United States)

    Szarka, Mate; Guttman, Andras

    2017-10-17

    We present the application of a smartphone anatomy based technology in the field of liquid phase bioseparations, particularly in capillary electrophoresis. A simple capillary electrophoresis system was built with LED induced fluorescence detection and a credit card sized minicomputer to prove the concept of real time fluorescent imaging (zone adjustable time-lapse fluorescence image processor) and separation controller. The system was evaluated by analyzing under- and overloaded aminopyrenetrisulfonate (APTS)-labeled oligosaccharide samples. The open source software based image processing tool allowed undistorted signal modulation (reprocessing) if the signal was inappropriate for the actual detection system settings (too low or too high). The novel smart detection tool for fluorescently labeled biomolecules greatly expands dynamic range and enables retrospective correction for injections with unsuitable signal levels without the necessity to repeat the analysis.

  13. Sources of variability among replicate samples separated by two-dimensional gel electrophoresis.

    Science.gov (United States)

    Bland, Alison M; Janech, Michael G; Almeida, Jonas S; Arthur, John M

    2010-04-01

    Two-dimensional gel electrophoresis (2DE) offers high-resolution separation for intact proteins. However, variability in the appearance of spots can limit the ability to identify true differences between conditions. Variability can occur at a number of levels. Individual samples can differ because of biological variability. Technical variability can occur during protein extraction, processing, or storage. Another potential source of variability occurs during analysis of the gels and is not a result of any of the causes of variability named above. We performed a study designed to focus only on the variability caused by analysis. We separated three aliquots of rat left ventricle and analyzed differences in protein abundance on the replicate 2D gels. As the samples loaded on each gel were identical, differences in protein abundance are caused by variability in separation or interpretation of the gels. Protein spots were compared across gels by quantile values to determine differences. Fourteen percent of spots had a maximum difference in intensity of 0.4 quantile values or more between replicates. We then looked individually at the spots to determine the cause of differences between the measured intensities. Reasons for differences were: failure to identify a spot (59%), differences in spot boundaries (13%), difference in the peak height (6%), and a combination of these factors (21). This study demonstrates that spot identification and characterization make major contributions to variability seen with 2DE. Methods to highlight why measured protein spot abundance is different could reduce these errors.

  14. Determination of five nitroimidazole residues in artificial porcine muscle tissue samples by capillary electrophoresis.

    Science.gov (United States)

    Lin, Yingyun; Su, Yan; Liao, Xiulin; Yang, Na; Yang, Xiupei; Choi, Martin M F

    2012-01-15

    A capillary electrophoresis (CE) method with ultraviolet detection has been developed for simultaneous detection and quantification of five nitroimidazoles including benzoylmetronidazole, dimetridazole, metronidazole, ronidazole, and secnidazole in porcine muscles. Nitroimidazoles in samples were extracted by ethyl acetate with subsequent clean-up by a strong cation exchange solid phase extraction column. The clean extracts were subjected to CE separation with optimal experimental conditions: pH 3.0 running buffer containing 25mM sodium phosphate and 0.10mM tetrabutylammonium bromide, 5s hydrodynamic injection at 0.5psi and 28kV separation voltage. The nitroimidazoles could be monitored and detected at 320nm within 18min. The limits of detection were below 1.0μg/kg and limits of quantification were lower than 3.2μg/kg for all nitroimidazoles in the muscle samples. The recoveries and relative standard deviations were 85.4-96.0, 83.5-92.5, 1.3-3.9, and 1.1-4.2%, respectively for the intra-day and inter-day analyses. The proposed CE method has been successfully applied to determine nitroimidazoles in artificial porcine muscle samples with good accuracy and recovery, demonstrating that it has potential for detection and quantification of multi-nitroimidazole residue in real muscle samples. Copyright © 2011 Elsevier B.V. All rights reserved.

  15. [Determination of arsenic speciation in Scomberomorus niphonius by capillary electrophoresis-inductively coupled plasma mass spectrometry].

    Science.gov (United States)

    Chen, Fa-rong; Zheng, Li; Wang, Zhi-Guang; Sun, Jie; Han, Li-Hui; Wang, Xiao-ru

    2014-06-01

    A method for the detection of arsenocholine (AsC), arsenobetaine (AsB), As(III), dimethylarsinic (DMA), monomethylarsonic (MMA) and As (V) by capillary electrophoresis-inductively coupled plasma mass spectrometry (CE-ICP-MS) was established. The results showed that the six species of arsenic were separated within 20 min under the optimized conditions. Good linearities of 6 arsenic species were observed in the range from 2 to 50 μg x L(-1) with the linear correlation greater than 0.996, the detection limits were 0.10-1.08 μg x L(-1) and the RSDs (n = 5) of the peak areas were smaller than 7%. The method was successfully adopted to the determination of the species in Scomberomorus niphonius. The recoveries were between 93% and 98%, and we found the arsenobetaine (AsB) was the main species in the sample. The method was suitable for the analysis of other biological samples with the advantages of good stability, less sample consumption, short analysis time and convenience.

  16. Selectivity in capillary electrophoresis:  application to chiral separations with cyclodextrins.

    Science.gov (United States)

    Lelièvre, F; Gareil, P; Jardy, A

    1997-02-01

    In order to accurately evaluate the performances of any electrolyte medium, a clear concept of selectivity in capillary electrophoresis and related electroseparation techniques is proposed. Selectivity is defined as the ratio of the affinity factors of both analytes for a separating agent (phase, pseudophase, or complexing agent present in the background electrolyte). When in the presence of a complexing agent and if only 1:1 complexation occurs, selectivity corresponds to the ratio of the apparent binding constants and is independent of the concentration of the complexing agent. This concept is illustrated through the separations of neutral and anionic enantiomers in the presence of a cationic cyclodextrin, the mono(6-amino-6-deoxy)-β-cyclodextrin, as a chiral complexing agent. The values obtained for different pairs of enantiomers are discussed with regard to the functional groups that distinguish them. When the analytes have the same mobilities in free solution and in their complexed form, then the resolution equation developed in micellar electrokinetic chromatography may be applied and optimum conditions (affinity factors, chiral agent concentration) can be predicted.

  17. Enantioselective analysis of fluoxetine in pharmaceutical formulations by capillary zone electrophoresis

    Directory of Open Access Journals (Sweden)

    Melania Cârcu-Dobrin

    2017-03-01

    Full Text Available Fluoxetine is an antidepressant, a selective serotonin reuptake inhibitor (SSRI used primarily in the treatment of major depression, panic disorder and obsessive compulsive disorder. Chiral separation of racemic fluoxetine is necessary due to its enantioselective metabolism. In order to develop a suitable method for chiral separation of fluoxetine, cyclodextrin (CD modified capillary electrophoresis (CE was employed. A large number of native and derivatized, neutral and ionized CD derivatives were screened to find the optimal chiral selector. As a result of this process, heptakis(2,3,6-tri-O-methyl-β-CD (TRIMEB was selected for enantiomeric discrimination. A factorial analysis study was performed by orthogonal experimental design in which several factors are varied at the same time to optimize the separation method. The optimized method (50 mM phosphate buffer, pH = 5.0, 10 mM TRIMEB, 15 °C, + 20 kV, 50 mbar/1 s, detection at 230 nm was successful for baseline separation of fluoxetine enantiomers within 5 min. Our method was validated according to ICH guidelines and proved to be sensitive, linear, accurate and precise for the chiral separation of fluoxetine.

  18. Analysis of O-Glycopeptides by Acetone Enrichment and Capillary Electrophoresis-Mass Spectrometry.

    Science.gov (United States)

    Mancera-Arteu, Montserrat; Giménez, Estela; Benavente, Fernando; Barbosa, José; Sanz-Nebot, Victòria

    2017-11-03

    Acetone precipitation was evaluated as a rapid, simple, low-cost, and efficient method for the selective purification of O-glycopeptides from enzymatic digests of glycoproteins. Ovalbumin (OVA), human and bovine α 1 -acid glycoprotein (hAGP and bAGP), human apolipoprotein C-III (APO-C3), and recombinant human erythropoietin (rhEPO) were used to obtain enzymatic digests with a broad and varied set of peptides, N-glycopeptides, and O-glycopeptides. After digestion and before capillary electrophoresis mass spectrometry (CE-MS) analysis, the amount of ice-cold acetone added to the digests was optimized to maximize recoveries of O-glycopeptides. Furthermore, the different behavior of peptides, N- and O-glycopeptides was explained by studying with multivariate data analysis methods the influence of several physicochemical parameters and properties related to their composition and structure. Principal component analysis (PCA) and, afterward, partial least-squares discriminant analysis (PLS-DA) were used to identify the most significant variables and their importance to differentiate between peptides, N-glycopeptides and O-glycopeptides, or within these classes. This information was useful to understand precipitation of these compounds after addition of acetone and for the selection of the optimal conditions for purification of specific O-glycopeptide biomarkers. Special attention was paid to O 126 -glycopeptide glycoforms of rhEPO because of their applicability in biopharmaceutical quality control and doping analysis.

  19. Metabolomics, peptidomics and proteomics applications of capillary electrophoresis-mass spectrometry in Foodomics: A review

    International Nuclear Information System (INIS)

    Ibáñez, Clara; Simó, Carolina; García-Cañas, Virginia; Cifuentes, Alejandro; Castro-Puyana, María

    2013-01-01

    Graphical abstract: -- Highlights: •Foodomics allows studying food and nutrition through the application of advanced omics approaches. •CE-MS plays a crucial role as analytical platform to carry out omics studies. •CE-MS applications for food metabolomics, proteomics and peptidomics are presented. -- Abstract: In the current post-genomic era, Foodomics has been defined as a discipline that studies food and nutrition through the application of advanced omics approaches. Foodomics involves the use of genomics, transcriptomics, epigenetics, proteomics, peptidomics, and/or metabolomics to investigate food quality, safety, traceability and bioactivity. In this context, capillary electrophoresis-mass spectrometry (CE-MS) has been applied mainly in food proteomics, peptidomics and metabolomics. The aim of this review work is to present an overview of the most recent developments and applications of CE-MS as analytical platform for Foodomics, covering the relevant works published from 2008 to 2012. The review provides also information about the integration of several omics approaches in the new Foodomics field

  20. Reverse polarity capillary zone electrophoresis analysis of nitrate and nitrite in natural water samples

    Energy Technology Data Exchange (ETDEWEB)

    Metcalf, S.G.

    1998-06-11

    This paper describes the application of reverse polarity capillary zone electrophoresis (RPCE) for rapid and accurate determination of nitrate and nitrite in natural water samples. Using hexamethonium bromide (HMB) as an electroosmotic flow modifier in a borate buffer at pH 9.2, the resolution of nitrate and nitrite was accomplished in less than 3 minutes. RPCE was compared with ion chromatographic (IC) and cadmium reduction flow injection analysis (Cd-FIA) methods which are the two most commonly used standard methods for the analysis of natural water samples for nitrate and nitrite. When compared with the ion chromatographic method for the determination of nitrate and nitrite, RPCE reduced analysis time, decreased detection limits by a factor of 10, cut laboratory wastes by more than two orders of magnitude, and eliminated interferences commonly associated with IC. When compared with the cadmium reduction method, RPCE had the advantage of simultaneous determination of nitrate and nitrite, could be used in the presence of various metallic ions that normally interfere in cadmium reduction, and decreased detection limits by a factor of 10.

  1. Capillary electrophoresis method with UV-detection for analysis of free amino acids concentrations in food.

    Science.gov (United States)

    Omar, Mei Musa Ali; Elbashir, Abdalla Ahmed; Schmitz, Oliver J

    2017-01-01

    Simple and inexpensive capillary electrophoresis with UV-detection method (CE-UV) was optimized and validated for determination of six amino acids namely (alanine, asparagine, glutamine, proline, serine and valine) for Sudanese food. Amino acids in the samples were derivatized with 4-chloro-7-nitro-2,1,3-benzoxadiazole (NBD-Cl) prior to CE-UV analysis. Labeling reaction conditions (100mM borate buffer at pH 8.5, labeling reaction time 60min, temperature 70°C and NBD-Cl concentration 40mM) were systematically investigated. The optimal conditions for the separation were 100mM borate buffer at pH 9.7 and detected at 475nm. The method was validated in terms of linearity, limit of detection (LOD), limit of quantification (LOQ), precision (repeatability) (RSD%) and accuracy (recovery). Good linearity was achieved for all amino acids (r(2)>0.9981) in the concentration range of 2.5-40mg/L. The LODs in the range of 0.32-0.56mg/L were obtained. Recoveries of amino acids ranging from 85% to 108%, (n=3) were obtained. The validated method was successfully applied for the determination of amino acids for Sudanese food samples. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. [Investigation on the interaction between pentadecafluorooctanoic acid and human serum albumin by capillary electrophoresis].

    Science.gov (United States)

    Gu, Yi; Guo, Ming; Lü, Da; Hou, Ping; Yin, Xinxin

    2018-01-08

    Capillary electrophoresis (CE) has been used to establish the analytical method of interaction between pentadecafluorooctanoic acid (PFOA) and human serum albumin (HSA). Under the physiological conditions, the interaction model of PFOA and HSA were constructed. Mobility method, plug-plug kinetic (PPK) method and simplified Hummel-Dreyer method were used to determine the interaction between derivatives and HSA. Non-linear regression, Scatchard equation and Klotz equation were adopted to obtain the interaction parameters. The results showed that all the three methods can be used to analyze the interaction of PFOA-HSA system. According to the interaction parameters, the most suitable CE method is simplified Hummel-Dreyer method while the most suitable theoretical equation is non-linear regression. The binding parameters indicated that the interaction of PFOA-HSA system has only one type of binding sites and the binding is stable. The research results have illustrated the interaction between HSA and PFOA, and provided a beneficial reference for in-depth research of the toxic mechanism of PFOA.

  3. Green sample preparation for liquid chromatography and capillary electrophoresis of anionic and cationic analytes.

    Science.gov (United States)

    Wuethrich, Alain; Haddad, Paul R; Quirino, Joselito P

    2015-04-21

    A sample preparation device for the simultaneous enrichment and separation of cationic and anionic analytes was designed and implemented in an eight-channel configuration. The device is based on the use of an electric field to transfer the analytes from a large volume of sample into small volumes of electrolyte that was suspended into two glass micropipettes using a conductive hydrogel. This simple, economical, fast, and green (no organic solvent required) sample preparation scheme was evaluated using cationic and anionic herbicides as test analytes in water. The analytical figures of merit and ecological aspects were evaluated against the state-of-the-art sample preparation, solid-phase extraction. A drastic reduction in both sample preparation time (94% faster) and resources (99% less consumables used) was observed. Finally, the technique in combination with high-performance liquid chromatography and capillary electrophoresis was applied to analysis of quaternary ammonium and phenoxypropionic acid herbicides in fortified river water as well as drinking water (at levels relevant to Australian guidelines). The presented sustainable sample preparation approach could easily be applied to other charged analytes or adopted by other laboratories.

  4. [Determination of inorganic ions in explosive residues by capillary zone electrophoresis].

    Science.gov (United States)

    Feng, Junhe; Guo, Baoyuan; Lin, Jin-Ming; Xu, Jianzhong; Zhou, Hong; Sun, Yuyou; Liu, Yao; Quan, Yangke; Lu, Xiaoming

    2008-11-01

    Five anions (chlorate, perchlorate, nitrate, nitrite, and sulfate) and two cations (ammonium and potassium) in explosive residues have been separated and determined by capillary zone electrophoresis (CZE) with indirect ultraviolet detection. The electrolyte buffer for the cation separation was 10 mmol/L pyridine (pH 4.5) -3 mmol/L 18-crown-6-ether. Ammonium and potassium ions were baseline separated in less than 2.6 min with the detection limits of 0.10 mg/L and 0.25 mg/L (S/N = 3), respectively. The electrolyte buffer for the anion separation consisted of 40 mmol/L boric acid-1.8 mmol/L potassium dichromate-2 mmol/L sodium tetraborate (pH 8.6), and tetramethyl ammonium hydroxide (TMAOH) was used as electroosmotic flow modifier. All five anions were well separated in less than 4.6 min with the detection limit range of 0.10 - 1.85 mg/L (S/N = 3). The method was successfully used in real sample investigations to confirm the type of explosives.

  5. Simultaneous determination of isoniazid and p-aminosalicylic acid by capillary electrophoresis using chemiluminescence detection.

    Science.gov (United States)

    Zhang, Xinfeng; Xuan, Yuelan; Sun, Aimin; Lv, Yi; Hou, Xiandeng

    2009-01-01

    It was found that isoniazid (ISO) or p-aminosalicylic acid (PAS) could enhance the chemiluminescence (CL) emission from Cu (II)-luminol-hydrogen peroxide system, and the increased chemiluminescence signals were proportional to their concentrations, respectively. Based on this phenomenon, a chemiluminescence method coupled to capillary electrophoresis (CE) was established for simultaneous determination of ISO and PAS. The CE conditions including running buffer and running voltage were investigated in detail. The effects of the pH of H(2)O(2) solution and the concentrations of luminol, H(2)O(2) and Cu (II) on the CL signal were also investigated carefully. Under the optimized conditions, the analysis could be accomplished within 10 min, with the limits of detection of 0.3 microg mL(-1) for ISO and 1.1 microg mL(-1) for PAS, corresponding to 7.2 and 26.4 pg per injection (24 nL), respectively. Finally, the method was validated by determining the two analytes in pharmaceutical preparation and spiked human serum samples. The results of pharmaceutical tablet analysis were in good agreement with the labeled amounts. The recoveries for ISO and PAS in human serum were in the range of 92-104% and 90-113%, respectively. Copyright 2008 John Wiley & Sons, Ltd.

  6. Effects of organic solvent and cationic additive on capillary electrophoresis of peptides

    International Nuclear Information System (INIS)

    Surugau, L.N.; Bergstrom, Ed T.

    2008-01-01

    Capillary electrophoresis (CE) of nine peptides namely, bradykinin, bradykinin fragment 1-5, substance P, Arg 8 -vasopressin, luteinizing hormone-releasing hormone (LHRH), bombesin, leucine-enkephalin, methionine-enkephalin and oxytocin were carried out using 0.5 % and 1.0 % formic acid (FA) as the separation buffers, added with acetonitrile (ACN) and triethylamine (TEA) as an additive at low pH. The electrophoretic behaviour of these peptides was examined at different concentration of TEA (0, 10, 20, 30, 40 and 50 mM), and ACN (30, 40, 50, 60, 70 %) at their respective measured final pH. The results showed that all nine peptides were fully resolved with addition of 10 - 20 mM TEA. Peak efficiency was improved significantly by increasing TEA concentration up to 40 mM where 800 000 m -1 was obtained. Without TEA, the closely related enkephalins were co-migrating. Interestingly, by addition of as little as 5 mM TEA has sufficient to separate them almost at baseline. Increasing ACN to 40 % has shortened the analysis time by ca. 1 min. However, further increase of ACN can cause peak broadening and current instability. (author)

  7. Effects of organic solvent and cationic additive on capillary electrophoresis of peptides

    International Nuclear Information System (INIS)

    Surugau, L.N.; Bergstrom, E.T.

    2008-01-01

    Capillary electrophoresis (CE) of nine peptides namely, bradykinin, bradykinin fragment 1-5, substance P, Arg 8 -vasopressin, luteinizing hormone-releasing hormone (LHRH), bombesin, leucine-enkephalin, methionine-enkephalin and oxytocin were carried out using 0.5 % and 1.0 % formic acid (FA) as the separation buffers, added with acetonitrile (ACN) and triethylamine (TEA) as an additive at low pH. The electrophoretic behavior of these peptides was examined at different concentration of TEA (0, 10, 20, 30, 40 and 50 mM), and ACN (30, 40, 50, 60, 70 %) at their respective measured final pH. The results showed that all nine peptides were fully resolved with addition of 10-20 mM TEA. Peak efficiency was improved significantly by increasing TEA concentration up to 40 mM where 800 000 m -1 was obtained. Without TEA, the closely related enkephalins were co-migrating. Interestingly, by addition of as little as 5 mM TEA has sufficient to separate them almost at baseline. Increasing ACN to 40 % has shortened the analysis time by ca. 1 min. However, further increase of ACN can cause peak broadening and current instability. (author)

  8. Development of a fraction collection approach in capillary electrophoresis SELEX for aptamer selection.

    Science.gov (United States)

    Luo, Zhaofeng; Zhou, Hongmin; Jiang, Hao; Ou, Huichao; Li, Xin; Zhang, Liyun

    2015-04-21

    Aptamers have attracted much attention due to their ability to bind to target molecules with high affinity and specificity. The development of an approach capable of efficiently generating aptamers through systematic evolution of ligands by exponential enrichment (SELEX) is particularly challenging. Herein, a fraction collection approach in capillary electrophoresis SELEX (FCE-SELEX) for the partition of a bound DNA-target complex is developed. By integrating fraction collection with a facile oil seal method for avoiding contamination while amplifying the bound DNA-target complex, in a single round of selection, a streptavidin-binding aptamer (SBA) has been generated. The affinity of aptamer SBA-36 for streptavidin (SA) is determined as 30.8 nM by surface plasmon resonance (SPR). Selectivity and biotin competition experiments demonstrate that the SBA-36 aptamer selected by FCE-SELEX is as efficient as those from other methods. Based on the ability of fraction collection in partition and collection of the aptamer-target complex from the original DNA library, FCE-SELEX can be a universal tool for the development of aptamers.

  9. Grapevine tissues and phenology differentially affect soluble carbohydrates determination by capillary electrophoresis.

    Science.gov (United States)

    Moreno, Daniela; Berli, Federico; Bottini, Rubén; Piccoli, Patricia N; Silva, María F

    2017-09-01

    Soluble carbohydrates distribution depends on plant physiology and, among other important factors, determines fruit yield and quality. In plant biology, the analysis of sugars is useful for many purposes, including metabolic studies. Capillary electrophoresis (CE) proved to be a powerful green separation technique with minimal sample preparation, even in complex plant tissues, that can provide high-resolution efficiency. Matrix effect refers to alterations in the analytical response caused by components of a sample other than the analyte of interest. Thus, the assessment and reduction of the matrix factor is fundamental for metabolic studies in different matrices. The present study evaluated the source and levels of matrix effects in the determination of most abundant sugars in grapevine tissues (mature and young leaves, berries and roots) at two phenological growth stages. Sucrose was the sugar that showed the least matrix effects, while fructose was the most affected analyte. Based on plant tissues, young leaves presented the smaller matrix effects, irrespectively of the phenology. These changes may be attributed to considerable differences at chemical composition of grapevine tissues with plant development. Therefore, matrix effect should be an important concern for plant metabolomics. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  10. Separation of thiosulfate and the polythionates in gold thiosulfate leach solutions by capillary electrophoresis.

    Science.gov (United States)

    O'Reilly, John W; Dicinoski, Greg W; Miura, Yasuyuki; Haddad, Paul R

    2003-06-01

    A technique for the separation of thiosulfate (S(2)O(3) (2-)), polythionates (S(x)O(6) (2-), x = 3 to 5) and the gold(I) thiosulfate complex (Au(S(2)O(3))(2) (3-)) using capillary electrophoresis with simultaneous UV detection at 195 and 214 nm is presented. The five species were separated in under 3 min with a total analysis time of 8 min, using an electrolyte containing 25 mM 2,2-bis(hydroxymethyl)-2,2',2"-nitrilotriethanol (bis-tris) adjusted to pH 6.0 with sulfuric acid and an applied voltage of -30 kV. While the gold(I) thiosulfate complex could be separated from the other analytes of interest under these conditions, the quantification of this complex was not possible due to inconsistent peak areas and peak splitting effects induced by the sulfur-oxygen species in the leach matrix. Detection limits calculated for 3s pressure injection at 50 mbar ranged between 0.5-2 microM. The method was linear over the ranges 40-8000, 10-2000, 10-2000, and 5-2000 microM for thiosulfate, trithionate, tetrathionate, and pentathionate, respectively. The technique was applied successfully to leach liquors containing 0.5 M ammonium thiosulfate, 2 M ammonia, 0.05 M copper sulfate and 20% w/v gold ore, diluted 1:100 prior to analysis.

  11. Speciation of organotin compounds by capillary electrophoresis: comparison of aqueous and mixed organic-aqueous systems

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Lei; Matysik, Frank-Michael; Glaeser, Petra [Universitaet Leipzig, Institut fuer Analytische Chemie, Leipzig (Germany)

    2004-10-01

    A capillary electrophoresis method with direct ultraviolet detection was developed for the analysis of organotin species. Despite the fact that direct detection of organotin compounds by ultraviolet absorption is difficult because most organotins possess poor chromophoric properties, the application of low wavelength ({lambda}=200 nm) and mixed organic-aqueous media enabled a significant enhancement in sensitivity. A mixed organic-aqueous system (10% methanol/40% acetonitrile/50% H{sub 2}O) containing acetic acid and tetrabutylammonium perchlorate formed the basis for rapid, efficient and sensitive determinations of organotin cations such as tripropyltin, tributyltin, triphenyltin and diphenyltin. The concentration limits of detection (LOD) for the four organotin compounds were in the range of 0.4-14 {mu}M, comparable to that obtained with the most sensitive indirect UV method reported until now, and took advantage of a stable baseline, a symmetric peak shape and an absence of disturbing system peaks. The relative standard deviations (n=7) for the relative peak time and peak area were 0.44-0.77 and 4.8-5.8%, respectively. In addition to sensitivity enhancements, the use of organic-aqueous systems instead of pure aqueous media resulted in improved selectivity and efficiency of separations. (orig.)

  12. Authentication of "Cereza del Jerte" sweet cherry varieties by free zone capillary electrophoresis (FZCE).

    Science.gov (United States)

    Serradilla, Manuel J; Martín, Alberto; Aranda, Emilio; Hernández, Alejandro; Benito, María J; Lopez-Corrales, Margarita; Córdoba, María de Guía

    2008-11-15

    The purpose of this work was to develop a procedure based on protein analysis by free zone capillary electrophoresis (FZCE) that can be used as an alternative to other methods in the determination of sweet cherry varieties for the authentication of "Cereza del Jerte". Two autochthonous varieties of sweet cherry type "Picota", 'Ambrunés' and 'Pico Negro', and the foreign variety 'Sweetheart' were used in the study. Two protocols for extracting the methanol-soluble proteins were tested. On the basis of the results, direct evaporation with nitrogen of a methanol extract was included in the extraction protocol for routine analysis. This method was found to give excellent repeatability of the corrected migration time (CMT), and showed greater effectiveness in discriminating sweet cherry varieties than the SDS-PAGE technique. Three peaks found in the FZCE electropherograms were investigated as a basis for discriminating between varieties. In addition, the FZCE analysis of methanol-soluble proteins provides information about the physico-chemical parameters relevant to the sensorial quality of the sweet cherries. Copyright © 2008 Elsevier Ltd. All rights reserved.

  13. Capillary electrophoresis with electrochemiluminescent detection for highly sensitive assay of genetically modified organisms.

    Science.gov (United States)

    Guo, Longhua; Yang, Huanghao; Qiu, Bin; Xiao, Xueyang; Xue, Linlin; Kim, Donghwan; Chen, Guonan

    2009-12-01

    A capillary electrophoresis coupled with electrochemiluminescent detection system (CE-ECL) was developed for the detection of polymerase chain reaction (PCR) amplicons. The ECL luminophore, tris(1,10-phenanthroline) ruthenium(II) (Ru(phen)(3)(2+)), was labeled to the PCR primers before amplification. Ru(phen)(3)(2+) was then introduced to PCR amplicons by PCR amplification. Eventually, the PCR amplicons were separated and detected by the homemade CE-ECL system. The detection of a typical genetically modified organism (GMO), Roundup Ready Soy (RRS), was shown as an example to demonstrate the reliability of the proposed approach. Four pairs of primers were amplified by multiple PCR (MPCR) simultaneously, three of which were targeted on the specific sequence of exogenous genes of RRS, and another was targeted on the endogenous reference gene of soybean. Both the conditions for PCR amplification and CE-ECL separation and detection were investigated in detail. Results showed that, under the optimal conditions, the proposed method can accurately identifying RRS. The corresponding limit of detection (LOD) was below 0.01% with 35 PCR cycles.

  14. Dual-signal amplification strategy for ultrasensitive chemiluminescence detection of PDGF-BB in capillary electrophoresis.

    Science.gov (United States)

    Cao, Jun-Tao; Wang, Hui; Ren, Shu-Wei; Chen, Yong-Hong; Liu, Yan-Ming

    2015-12-01

    Many efforts have been made toward the achievement of high sensitivity in capillary electrophoresis coupled with chemiluminescence detection (CE-CL). This work describes a novel dual-signal amplification strategy for highly specific and ultrasensitive CL detection of human platelet-derived growth factor-BB (PDGF-BB) using both aptamer and horseradish peroxidase (HRP) modified gold nanoparticles (HRP-AuNPs-aptamer) as nanoprobes in CE. Both AuNPs and HRP in the nanoprobes could amplify the CL signals in the luminol-H2 O2 CL system, owing to the excellent catalytic behavior of AuNPs and HRP in the CL system. Meanwhile, the high affinity of aptamer modified on the AuNPs allows detection with high specificity. As proof-of-concept, the proposed method was employed to quantify the concentration of PDGF-BB from 0.50 to 250 fm with a detection limit of 0.21 fm. The applicability of the assay was further demonstrated in the analysis of PDGF-BB in human serum samples with acceptable accuracy and reliability. The result of this study exhibits distinct advantages, such as high sensitivity, good specificity, simplicity, and very small sample consumption. The good performances of the proposed strategy provide a powerful avenue for ultrasensitive detection of rare proteins in biological sample, showing great promise in biochemical analysis. Copyright © 2015 John Wiley & Sons, Ltd.

  15. Fast Separation and Determination of Flavonoids in Honey Samples by Capillary Zone Electrophoresis

    Directory of Open Access Journals (Sweden)

    Jian-Qiu Tu

    2017-03-01

    Full Text Available Flavonoids have crucial applications in the biological and physiological fields. Honey, as an important sweet food made by bees, is rich in flavonoids. In this paper, the analytical method for flavonoids determination in different sorts of honey from different geographical locations was developed by capillary zone electrophoresis with direct ultraviolet detection. With a running buffer (borate, 20 mmol l−1 at pH of 8.4, four typical flavonoids, rutin, quercetin, kaempferol and myricetin, were separated in five minutes under a applied potential of 25 kV. A linear relationship within the range of 2.0 – 500 mg l−1 was found for these four kinds of flavonoids. Moreover, the detection limits ranged from 1.17 to 1.76 mg l−1. The recoveries lie in the range between 80 % – 107 %. The developed method was then used in the separation and determination of flavonoids in real honey samples collected from 12 geographical locations in the Henan Province of China. Rutin was detected in six, and quercetin in eight honey samples, which may be the markers for the identification of honey from different geographical origins.

  16. Separation of arsenic species by capillary electrophoresis with sample-stacking techniques

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Zu Liang; Naidu, Ravendra [Adelaide Laboratory, CSIRO Land and Water, PMB2, 5064, Glen Osmond, SA (Australia); Lin, Jin-Ming [Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, P.O. Box 2871, 100085, Beijing (China)

    2003-03-01

    A simple capillary zone electrophoresis procedure was developed for the separation of arsenic species (AsO{sub 2}{sup 2-}, AsO{sub 4}{sup 2-}, and dimethylarsinic acid, DMA). Both counter-electroosmotic and co-electroosmotic (EOF) modes were investigated for the separation of arsenic species with direct UV detection at 185 nm using 20 mmol L{sup -1} sodium phosphate as the electrolyte. The separation selectivity mainly depends on the separation modes and electrolyte pH. Inorganic anions (Cl{sup -}, NO{sub 2}{sup -}, NO{sub 3}{sup -} and SO{sub 4}{sup 2-}) presented in real samples did not interfere with arsenic speciation in either separation mode. To improve the detection limits, sample-stacking techniques, including large-volume sample stacking (LVSS) and field-amplified sample injection (FASI), were investigated for the preconcentration of As species in co-CZE mode. Less than 1 {mu}mol L{sup -1} of detection limits for As species were achieved using FASI. The proposed method was demonstrated for the separation and detection of As species in water. (orig.)

  17. Investigation of Interactions between Thrombin and Ten Phenolic Compounds by Affinity Capillary Electrophoresis and Molecular Docking

    Directory of Open Access Journals (Sweden)

    Qiao-Qiao Li

    2018-01-01

    Full Text Available Thrombin plays a vital role in blood coagulation, which is a key process involved in thrombosis by promoting platelet aggregation and converting fibrinogen to form the fibrin clot. In the receptor concept, drugs produce their therapeutic effects via interactions with the targets. Therefore, investigation of interaction between thrombin and small molecules is important to find out the potential thrombin inhibitor. In this study, affinity capillary electrophoresis (ACE and in silico molecular docking methods were developed to study the interaction between thrombin and ten phenolic compounds (p-hydroxybenzoic acid, protocatechuic acid, vanillic acid, gallic acid, catechin, epicatechin, dihydroquercetin, naringenin, apigenin, and baicalein. The ACE results showed that gallic acids and six flavonoid compounds had relative strong interactions with thrombin. In addition, the docking results indicated that all of optimal conformations of the six flavonoid compounds were positioned into the thrombin activity centre and had interaction with the HIS57 or SER195 which was the key residue to bind thrombin inhibitors such as argatroban. Herein, these six flavonoid compounds might have the potential of thrombin inhibition activity. In addition, the developed method in this study can be further applied to study the interactions of other molecules with thrombin.

  18. Review on the development of truly portable and in-situ capillary electrophoresis systems

    Science.gov (United States)

    Lewis, A. P.; Cranny, A.; Harris, N. R.; Green, N. G.; Wharton, J. A.; Wood, R. J. K.; Stokes, K. R.

    2013-04-01

    Capillary electrophoresis (CE) is a technique which uses an electric field to separate a mixed sample into its constituents. Portable CE systems enable this powerful analysis technique to be used in the field. Many of the challenges for portable systems are similar to those of autonomous in-situ analysis and therefore portable systems may be considered a stepping stone towards autonomous in-situ analysis. CE is widely used for biological and chemical analysis and example applications include: water quality analysis; drug development and quality control; proteomics and DNA analysis; counter-terrorism (explosive material identification) and corrosion monitoring. The technique is often limited to laboratory use, since it requires large electric fields, sensitive detection systems and fluidic control systems. All of these place restrictions in terms of: size, weight, cost, choice of operating solutions, choice of fabrication materials, electrical power and lifetime. In this review we bring together and critique the work by researchers addressing these issues. We emphasize the importance of a holistic approach for portable and in-situ CE systems and discuss all the aspects of the design. We identify gaps in the literature which require attention for the realization of both truly portable and in-situ CE systems.

  19. Capillary electrophoresis - Mass spectrometry metabolomics analysis revealed enrichment of hypotaurine in rat glioma tissues.

    Science.gov (United States)

    Gao, Peng; Ji, Min; Fang, Xueyan; Liu, Yingyang; Yu, Zhigang; Cao, Yunfeng; Sun, Aijun; Zhao, Liang; Zhang, Yong

    2017-11-15

    Glioma is one of the most lethal brain malignancies with unknown etiologies. Many metabolomics analysis aiming at diverse kinds of samples had been performed. Due to the varied adopted analytical platforms, the reported disease-related metabolites were not consistent across different studies. Comparable metabolomics results are more likely to be acquired by analyzing the same sample types with identical analytical platform. For tumor researches, tissue samples metabolomics analysis own the unique advantage that it can gain more direct insight into disease-specific pathological molecules. In this light, a previous reported capillary electrophoresis - mass spectrometry human tissues metabolomics analysis method was employed to profile the metabolome of rat C6 cell implantation gliomas and the corresponding precancerous tissues. It was found that 9 metabolites increased in the glioma tissues. Of them, hypotaurine was the only metabolite that enriched in the malignant tissues as what had been reported in the relevant human tissues metabolomics analysis. Furthermore, hypotaurine was also proved to inhibit α-ketoglutarate-dependent dioxygenases (2-KDDs) through immunocytochemistry staining and in vitro enzymatic activity assays by using C6 cell cultures. This study reinforced the previous conclusion that hypotaurine acted as a competitive inhibitor of 2-KDDs and proved the value of metabolomics in oncology studies. Copyright © 2017. Published by Elsevier Inc.

  20. Methoxypropylamino β-cyclodextrin clicked AC regioisomer for enantioseparations in capillary electrophoresis

    International Nuclear Information System (INIS)

    Zhou, Jie; Wang, Yiying; Liu, Yun; Tang, Jian; Tang, Weihua

    2015-01-01

    Highlights: In this paper, we demonstrate: • The click synthesis of a AC regioisomer cationic cyclodextrin (CD) as chiral selector. • The good enantioselectivities (chiral resolution over 5) for acidic racemates. • The strong chiral recognition of new CD by NMR study. • Baseline enantioseparation of some acidic racemates at CD of 0.5 mM. - Abstract: In this work, a novel methoxypropylamino β-cyclodextrin (β-CD) clicked AC regioisomer, 6 A -4-hydroxyethyl-1,2,3-triazolyl-6 C -3-methoxypropylamino β-cyclodextrin (HETz-MPrAMCD), was synthesized via nucleophilic addition and click chemistry. The chiral separation ability of this AC regioisomer cationic CD was evaluated toward 7 ampholytic and 13 acidic racemates by capillary electrophoresis. Dependence of enantioselectivity and resolution on buffer pH (5.5–8.0) and chiral selector concentration (0.5–7.5 mM) was investigated. Enantioselectivities (α ≥ 1.05) could be achieved for most analytes under optimal conditions except dansyl-DL-noreleucine and dansyl-DL-serine. The highest resolutions for 2-chloromandelic acid p-hydroxymandelic acid were 15.6 and 9.7 respectively. The inclusion complexation between HETz-MPrAMCD and each 3-phenyllactic acid enantiomer was also revealed with nuclear magnetic resonance study

  1. Methoxypropylamino β-cyclodextrin clicked AC regioisomer for enantioseparations in capillary electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Jie; Wang, Yiying; Liu, Yun; Tang, Jian; Tang, Weihua, E-mail: whtang@mail.njust.edu.cn

    2015-04-08

    Highlights: In this paper, we demonstrate: • The click synthesis of a AC regioisomer cationic cyclodextrin (CD) as chiral selector. • The good enantioselectivities (chiral resolution over 5) for acidic racemates. • The strong chiral recognition of new CD by NMR study. • Baseline enantioseparation of some acidic racemates at CD of 0.5 mM. - Abstract: In this work, a novel methoxypropylamino β-cyclodextrin (β-CD) clicked AC regioisomer, 6{sup A}-4-hydroxyethyl-1,2,3-triazolyl-6{sup C}-3-methoxypropylamino β-cyclodextrin (HETz-MPrAMCD), was synthesized via nucleophilic addition and click chemistry. The chiral separation ability of this AC regioisomer cationic CD was evaluated toward 7 ampholytic and 13 acidic racemates by capillary electrophoresis. Dependence of enantioselectivity and resolution on buffer pH (5.5–8.0) and chiral selector concentration (0.5–7.5 mM) was investigated. Enantioselectivities (α ≥ 1.05) could be achieved for most analytes under optimal conditions except dansyl-DL-noreleucine and dansyl-DL-serine. The highest resolutions for 2-chloromandelic acid p-hydroxymandelic acid were 15.6 and 9.7 respectively. The inclusion complexation between HETz-MPrAMCD and each 3-phenyllactic acid enantiomer was also revealed with nuclear magnetic resonance study.

  2. Development of a fast capillary electrophoresis method for determination of carbohydrates in honey samples.

    Science.gov (United States)

    Rizelio, Viviane Maria; Tenfen, Laura; da Silveira, Roberta; Gonzaga, Luciano Valdemiro; Costa, Ana Carolina Oliveira; Fett, Roseane

    2012-05-15

    In this study, the determination of fructose, glucose and sucrose by capillary electrophoresis (CE) was investigated. The tendency of the analyte to undergo electromigration dispersion and the buffer capacity were evaluated using the Peakmaster(®) software and considered in the optimization of the background electrolyte, which was composed of 20 mmol L(-1) sorbic acid, 0.2 mmol L(-1) CTAB and 40 mmol L(-1) NaOH at pH 12.2. Under optimal CE conditions, the separation of the substances investigated was achieved in less than 2 min. The detection limits for the three analytes were in the range of 0.022 and 0.029 g L(-1) and precision measurements within 0.62-4.69% were achieved. The proposed methodology was applied in the quantitative analysis by direct injection of in honey samples to determine the main sugars presents. The samples were previously dissolved in deionized water and filtered with no other sample treatment. The mean values for fructose, glucose and sucrose were in the ranges of 33.65-45.46 g 100g(-1), 24.63-35.06 g 100g(-1) and <0.22-1.32 g 100g(-1), respectively. The good analytical performance of the method makes it suitable for implementation in food laboratories for the routine analysis of honey samples. Copyright © 2012 Elsevier B.V. All rights reserved.

  3. Review on the development of truly portable and in-situ capillary electrophoresis systems

    International Nuclear Information System (INIS)

    Lewis, A P; Cranny, A; Harris, N R; Green, N G; Wharton, J A; Wood, R J K; Stokes, K R

    2013-01-01

    Capillary electrophoresis (CE) is a technique which uses an electric field to separate a mixed sample into its constituents. Portable CE systems enable this powerful analysis technique to be used in the field. Many of the challenges for portable systems are similar to those of autonomous in-situ analysis and therefore portable systems may be considered a stepping stone towards autonomous in-situ analysis. CE is widely used for biological and chemical analysis and example applications include: water quality analysis; drug development and quality control; proteomics and DNA analysis; counter-terrorism (explosive material identification) and corrosion monitoring. The technique is often limited to laboratory use, since it requires large electric fields, sensitive detection systems and fluidic control systems. All of these place restrictions in terms of: size, weight, cost, choice of operating solutions, choice of fabrication materials, electrical power and lifetime. In this review we bring together and critique the work by researchers addressing these issues. We emphasize the importance of a holistic approach for portable and in-situ CE systems and discuss all the aspects of the design. We identify gaps in the literature which require attention for the realization of both truly portable and in-situ CE systems. (topical review)

  4. Simultaneous analysis of saturated and unsaturated fatty acids present in pequi fruits by capillary electrophoresis

    Directory of Open Access Journals (Sweden)

    Patrícia M. de Castro Barra

    2013-01-01

    Full Text Available In the current study, an alternative method has been proposed for simultaneous analysis of palmitic, stearic, oleic, linoleic, and linolenic acids by capillary zone electrophoresis (CZE using indirect detection. The background electrolyte (BGE used for the analysis of these fatty acids (FAs consisted of 15.0 mmol L−1 NaH2PO4/Na2HPO4 at pH 6.86, 4.0 mmol L−1 SDBS, 8.3 mmol L−1 Brij 35, 45% v/v acetonitrile (can, and 2.1% n-octanol. The FAs quantification of FAs was performed using a response factor approach, which provided a high analytical throughput for the real sample. The CZE method, which was applied successfully for the analysis of pequi pulp, has advantages such as short analysis time, absence of lipid fraction extraction and derivatization steps, and no significant difference in the 95% confidence intervals for FA quantification results, compared to the gas chromatography official method (AOCS Ce 1h-05.

  5. [Determination of penicillin intermediate and three penicillins in milk by high performance capillary electrophoresis].

    Science.gov (United States)

    Tian, Chunqiu; Tan, Huarong; Gao, Liping; Shen, Huqin; Qi, Kezong

    2011-11-01

    A high performance capillary electrophoresis (HPCE) method was developed for the simultaneous determination of penicillin intermediate and penicillins in milk, including 6-amino-penicillanic acid (6-APA), penicillin G (PEN), ampicillin (AMP) and amoxicillin (AMO). The main parameters including the ion concentration and pH value of running buffer, separation voltage and column temperature were optimized systematically by orthogonal test. The four penicillins (PENs) were baseline separated within 4.5 min with the running buffer of 40 mmol/L potassium dihydrogen phosphate-20 mmol/L borax solution (pH 7.8), separation voltage of 28 kV and column temperature of 30 degrees C. The calibration curves showed good linearity in the range of 1.56 - 100 mg/L, and the correlation coefficients (r2) were between 0.9979 and 0.9998. The average recoveries at three spiked levels were in the range of 84.91% - 96.72% with acceptable relative standard deviations (RSDs) of 1.11% - 9.11%. The method is simple, fast, accurate and suitable for the determination of penicillins in real samples.

  6. Genotyping of K-ras codons 12 and 13 mutations in colorectal cancer by capillary electrophoresis.

    Science.gov (United States)

    Chen, Yen-Ling; Chang, Ya-Sian; Chang, Jan-Gowth; Wu, Shou-Mei

    2009-06-26

    Point mutations of the K-ras gene located in codons 12 and 13 cause poor responses to the anti-epidermal growth factor receptor (anti-EGFR) therapy of colorectal cancer (CRC) patients. Besides, mutations of K-ras gene have also been proven to play an important role in human tumor progression. We established a simple and effective capillary electrophoresis (CE) method for simultaneous point mutation detection in codons 12 and 13 of K-ras gene. We combined one universal fluorescence-based nonhuman-sequence primer and two fragment-oriented primers in one tube, and performed this two-in-one polymerase chain reaction (PCR). PCR fragments included wild type and seven point mutations at codons 12 and 13 of K-ras gene. The amplicons were analyzed by single-strand conformation polymorphism (SSCP)-CE method. The CE analysis was performed by using a 1x Tris-borate-EDTA (TBE) buffer containing 1.5% (w/v) hydroxyethylcellulose (HEC) (MW 250,000) under reverse polarity with 15 degrees C and 30 degrees C. Ninety colorectal cancer patients were blindly genotyped using this developed method. The results showed good agreement with those of DNA sequencing method. The SSCP-CE was feasible for mutation screening of K-ras gene in populations.

  7. Determination of gentisin, isogentisin, and amarogentin in Gentiana lutea L. by capillary electrophoresis.

    Science.gov (United States)

    Citová, Ivana; Ganzera, Markus; Stuppner, Hermann; Solich, Petr

    2008-01-01

    A novel, fast, and simple capillary electrophoresis method has been developed for the analysis of gentisin, isogentisin, and amarogentin in roots of Gentiana lutea (yellow gentian), an herb traditionally used as gastric stimulant. Gentisin and isogentisin are xanthones showing potent inhibition of monoamine oxidase type A and B, amarogentin represents one of the bitter principles of Gentiana, responsible for its gastric-roborant effects. Optimal CE-separation conditions comprise a 100 mM sodium tetraborate buffer of pH 9.3, containing 10 mM beta-cyclodextrin as additive; optimum temperature and applied voltage were found to be 30 degrees C and 25 kV, respectively. Direct diode array detection at 260 nm (gentisin, isogentisin) and 242 nm (amarogentin) was performed, and the required analysis time was only 11 min. The developed method was validated for linearity, sensitivity, precision, and accuracy, and utilized to assay several commercially available G. lutea samples. Quantitative data obtained with the developed CE method are compared with HPLC results, and the advantages of each approach are discussed.

  8. Analysis of aromatic aldehydes in brandy and wine by high-performance capillary electrophoresis.

    Science.gov (United States)

    Panossian, A; Mamikonyan, G; Torosyan, M; Gabrielyan, E; Mkhitaryan, S

    2001-09-01

    A new method of analysis of vanillin, syringaldehyde, coniferaldehyde, and sinapaldehyde in brandy and wine by high-performance capillary electrophoresis is described. Electrophoretic mobility of these compounds is achieved by a borate buffer at pH 9.3. At this pH, the sensitivity of UV detection of these phenolic aldehydes also increases. UV absorptions at 348, 362, 404, and 422 nm were selected for monitoring vanillin, syringaldehyde, coniferaldehyde, and sinapaldehyde, respectively. This procedure was performed simultaneously during one run using a diode array detector. Samples of brandy or wine were analyzed directly without concentration, extraction, or any other preliminary treatment of the test sample. The limits of detection were found to be 0.275, 0.1425, 0.1475, and 0.1975 ppm for syringaldehyde, coniferaldehyde, sinapaldehyde, and vanillin, respectively, which is acceptable for analysis of both brandy and wine aged in oak barrels. The method has been shown to be linear in a range from 0.3 to 57 mg/L. Recoveries ranged between 99.9% and 107.7% for all of the compounds tested. Repeatability and reproducibility of the method were high. The relative standard deviation was consequently approximately 3% and also between 4.47% and 6.89% for all tested compounds. The method is useful for the identification of counterfeit brandy, which is easy to recognize by the absence of sinapaldehyde, syringaldehyde, and coniferaldehyde, which are not detectable in false brandy.

  9. A rapid sample screening method for authenticity control of whiskey using capillary electrophoresis with online preconcentration.

    Science.gov (United States)

    Heller, Melina; Vitali, Luciano; Oliveira, Marcone Augusto Leal; Costa, Ana Carolina O; Micke, Gustavo Amadeu

    2011-07-13

    The present study aimed to develop a methodology using capillary electrophoresis for the determination of sinapaldehyde, syringaldehyde, coniferaldehyde, and vanillin in whiskey samples. The main objective was to obtain a screening method to differentiate authentic samples from seized samples suspected of being false using the phenolic aldehydes as chemical markers. The optimized background electrolyte was composed of 20 mmol L(-1) sodium tetraborate with 10% MeOH at pH 9.3. The study examined two kinds of sample stacking, using a long-end injection mode: normal sample stacking (NSM) and sample stacking with matrix removal (SWMR). In SWMR, the optimized injection time of the samples was 42 s (SWMR42); at this time, no matrix effects were observed. Values of r were >0.99 for the both methods. The LOD and LOQ were better than 100 and 330 mg mL(-1) for NSM and better than 22 and 73 mg L(-1) for SWMR. The CE-UV reliability in the aldehyde analysis in the real sample was compared statistically with LC-MS/MS methodology, and no significant differences were found, with a 95% confidence interval between the methodologies.

  10. Reverse polarity capillary zone electrophoresis analysis of nitrate and nitrite in natural water samples

    International Nuclear Information System (INIS)

    Metcalf, S.G.

    1998-01-01

    This paper describes the application of reverse polarity capillary zone electrophoresis (RPCE) for rapid and accurate determination of nitrate and nitrite in natural water samples. Using hexamethonium bromide (HMB) as an electroosmotic flow modifier in a borate buffer at pH 9.2, the resolution of nitrate and nitrite was accomplished in less than 3 minutes. RPCE was compared with ion chromatographic (IC) and cadmium reduction flow injection analysis (Cd-FIA) methods which are the two most commonly used standard methods for the analysis of natural water samples for nitrate and nitrite. When compared with the ion chromatographic method for the determination of nitrate and nitrite, RPCE reduced analysis time, decreased detection limits by a factor of 10, cut laboratory wastes by more than two orders of magnitude, and eliminated interferences commonly associated with IC. When compared with the cadmium reduction method, RPCE had the advantage of simultaneous determination of nitrate and nitrite, could be used in the presence of various metallic ions that normally interfere in cadmium reduction, and decreased detection limits by a factor of 10

  11. Analysis of Three Compounds in Flos Farfarae by Capillary Electrophoresis with Large-Volume Sample Stacking

    Directory of Open Access Journals (Sweden)

    Hai-xia Yu

    2017-01-01

    Full Text Available The aim of this study was to develop a method combining an online concentration and high-efficiency capillary electrophoresis separation to analyze and detect three compounds (rutin, hyperoside, and chlorogenic acid in Flos Farfarae. In order to get good resolution and enrichment, several parameters such as the choice of running buffer, pH and concentration of the running buffer, organic modifier, temperature, and separation voltage were all investigated. The optimized conditions were obtained as follows: the buffer of 40 mM NaH2P04-40 mM Borax-30% v/v methanol (pH 9.0; the sample hydrodynamic injection of up to 4 s at 0.5 psi; 20 kV applied voltage. The diode-array detector was used, and the detection wavelength was 364 nm. Based on peak area, higher levels of selective and sensitive improvements in analysis were observed and about 14-, 26-, and 5-fold enrichment of rutin, hyperoside, and chlorogenic acid were achieved, respectively. This method was successfully applied to determine the three compounds in Flos Farfarae. The linear curve of peak response versus concentration was from 20 to 400 µg/ml, 16.5 to 330 µg/mL, and 25 to 500 µg/mL, respectively. The regression coefficients were 0.9998, 0.9999, and 0.9991, respectively.

  12. Distinction of water-soluble constituents between natural and cultured Cordyceps by capillary electrophoresis.

    Science.gov (United States)

    Li, S P; Song, Z H; Dong, T T X; Ji, Z N; Lo, C K; Zhu, S Q; Tsim, K W K

    2004-11-01

    Cordyceps is an expensive traditional Chinese medicine, which has anti-tumor activity and significant effects on the immune system. In Southeast Asia, Cordyceps is commonly sold in capsule form as a health food product. Most of these products are derived from cultured Cordyceps mycelia. Because of the price difference, some manufacturers claim their products are from natural Cordyceps. In order to distinguish among various types of Cordyceps in the market, the profiles of water-soluble constituents derived from different sources of Cordyceps were determined by capillary electrophoresis (CE). Both natural and cultured Cordyceps showed three peak clusters migrated at 5-7, 9-11 and 12-13 min, and the height and resolution of these peak clusters were rather distinct. Peak cluster at 9-11 min was identified as adenosine, guanosine and uridine, and shared a similarity between natural and cultured products. In contrast, the peak cluster at 5-7 min was characteristic of natural Cordyceps, regardless of hosts and sources. By using the peak characteristics of CE profiles of different Cordyceps samples, hierarchical clustering analysis was performed. The result shows that those samples of natural Cordyceps were grouped together distinct from the cultured and commercial products. Thus, the CE profiles could serve as fingerprints for the quality control of Cordyceps.

  13. Simultaneous Detection of Genetically Modified Organisms in a Mixture by Multiplex PCR-Chip Capillary Electrophoresis.

    Science.gov (United States)

    Patwardhan, Supriya; Dasari, Srikanth; Bhagavatula, Krishna; Mueller, Steffen; Deepak, Saligrama Adavigowda; Ghosh, Sudip; Basak, Sanjay

    2015-01-01

    An efficient PCR-based method to trace genetically modified food and feed products is in demand due to regulatory requirements and contaminant issues in India. However, post-PCR detection with conventional methods has limited sensitivity in amplicon separation that is crucial in multiplexing. The study aimed to develop a sensitive post-PCR detection method by using PCR-chip capillary electrophoresis (PCR-CCE) to detect and identify specific genetically modified organisms in their genomic DNA mixture by targeting event-specific nucleotide sequences. Using the PCR-CCE approach, novel multiplex methods were developed to detect MON531 cotton, EH 92-527-1 potato, Bt176 maize, GT73 canola, or GA21 maize simultaneously when their genomic DNAs in mixtures were amplified using their primer mixture. The repeatability RSD (RSDr) of the peak migration time was 0.06 and 3.88% for the MON531 and Bt176, respectively. The RSD (RSDR) of the Cry1Ac peak ranged from 0.12 to 0.40% in multiplex methods. The method was sensitive in resolving amplicon of size difference up to 4 bp. The PCR-CCE method is suitable to detect multiple genetically modified events in a composite DNA sample by tagging their event specific sequences.

  14. Electrohydrodynamic properties of succinoglycan as probed by fluorescence correlation spectroscopy, potentiometric titration and capillary electrophoresis.

    Science.gov (United States)

    Duval, Jérôme F L; Slaveykova, Vera I; Hosse, Monika; Buffle, Jacques; Wilkinson, Kevin J

    2006-10-01

    The electrostatic, hydrodynamic and conformational properties of aqueous solutions of succinoglycan have been analyzed by fluorescence correlation spectroscopy (FCS), proton titration, and capillary electrophoresis (CE) over a large range of pH values and electrolyte (NaCl) concentrations. Using the theoretical formalism developed previously for the electrokinetic properties of soft, permeable particles, a quantitative analysis for the electro-hydrodynamics of succinoglycan is performed by taking into account, in a self-consistent manner, the measured values of the diffusion coefficients, electric charge densities, and electrophoretic mobilities. For that purpose, two limiting conformations for the polysaccharide in solution are tested, i.e. succinoglycan behaves as (i) a spherical, random coil polymer or (ii) a rodlike particle with charged lateral chains. The results show that satisfactory modeling of the titration data for ionic strengths larger than 50 mM can be accomplished using both geometries over the entire range of pH values. Electrophoretic mobilities measured for sufficiently large pH values (pH > 5-6) are in line with predictions based on either model. The best manner to discriminate between these two conceptual models is briefly discussed. For low pH values (pH < 5), both models indicate aggregation, resulting in an increase of the hydrodynamic permeability and a decrease of the diffusion coefficient.

  15. A direct and rapid method to determine cyanide in urine by capillary electrophoresis.

    Science.gov (United States)

    Zhang, Qiyang; Maddukuri, Naveen; Gong, Maojun

    2015-10-02

    Cyanides are poisonous chemicals that widely exist in nature and industrial processes as well as accidental fires. Rapid and accurate determination of cyanide exposure would facilitate forensic investigation, medical diagnosis, and chronic cyanide monitoring. Here, a rapid and direct method was developed for the determination of cyanide ions in urinary samples. This technique was based on an integrated capillary electrophoresis system coupled with laser-induced fluorescence (LIF) detection. Cyanide ions were derivatized with naphthalene-2,3-dicarboxaldehyde (NDA) and a primary amine (glycine) for LIF detection. Three separate reagents, NDA, glycine, and cyanide sample, were mixed online, which secured uniform conditions between samples for cyanide derivatization and reduced the risk of precipitation formation of mixtures. Conditions were optimized; the derivatization was completed in 2-4min, and the separation was observed in 25s. The limit of detection (LOD) was 4.0nM at 3-fold signal-to-noise ratio for standard cyanide in buffer. The cyanide levels in urine samples from smokers and non-smokers were determined by using the method of standard addition, which demonstrated significant difference of cyanide levels in urinary samples from the two groups of people. The developed method was rapid and accurate, and is anticipated to be applicable to cyanide detection in waste water with appropriate modification. Published by Elsevier B.V.

  16. Quantitative analysis of flavonoids and phenolic acid in Coreopsis tinctoria Nutt. by capillary zone electrophoresis.

    Science.gov (United States)

    Deng, Yong; Lam, Shing-Chung; Zhao, Jing; Li, Shao-Ping

    2017-10-01

    Capillary zone electrophoresis was developed for the simultaneous determination of five flavonoids and one phenolic acid, including taxifolin-7-O-glucoside, flavanomarein, quercetagetin-7-O-glucoside, okanin 4'-O-glucoside, okanin, and chlorogenic acid, in different parts and origins of Coreopsis tinctoria and its related species. Effects of acidity, running-buffer concentration, and modifier concentration were investigated to determine the optimum conditions for analyte determination. Analysis was performed within 18 min by using 50 mM borax buffer containing 15% acetonitrile as a modifier (pH 9.0) at 25 kV and 25°C. Hyperoside was used as internal standard for quantification. The method was accurate, simple, and repeatable, and was successfully applied to the analysis in 13 samples with satisfactory assay results. Results showed that C. tinctoria obviously differed from the related flower tea materials, "Hangju" and "Gongju". The parts (flowers, buds, seeds, stems, and leaves) of C. tinctoria also varied among one another. This study can serve as a foundation for the quality control and pharmacological evaluation of different parts of C. tinctoria and its related species. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. Influence of Immobilized Biomolecules on Magnetic Bead Plug Formation and Retention in Capillary Electrophoresis

    Science.gov (United States)

    Henken, Rachel L.; Chantiwas, Rattikan; Gilman, S. Douglass

    2012-01-01

    Significant changes in the formation and retention of magnetic bead plugs in a capillary during electrophoresis were studied, and it was demonstrated that these effects were due to the type of biological molecule immobilized on the surface of these beads. Three biological molecules, an antibody, an oligonucleotide and alkaline phosphatase, were attached to otherwise identical streptavidin-coated magnetic beads through biotin-avidin binding in order to isolate differences in bead immobilization in a magnetic field resulting from the type of biological molecule immobilized on the bead surface. Alkaline phosphatase also was attached to the magnetic beads using epoxy groups on the bead surfaces (instead of avidin-biotin binding) to study the impact of immobilization chemistry. The formation and retention of magnetic bead plugs were studied quantitatively using light scattering detection of magnetic particles eluting from the bead plugs and qualitatively using microscopy. Both the type of biomolecule immobilized on the magnetic bead surface and the chemistry used to link the biomolecule to the magnetic bead impacted the formation and retention of the bead plugs. PMID:22437880

  18. Capillary zone electrophoresis-tandem mass spectrometry detects low concentration host cell impurities in monoclonal antibodies

    Science.gov (United States)

    Zhu, Guijie; Sun, Liangliang; Heidbrink-Thompson, Jennifer; Kuntumalla, Srilatha; Lin, Hung-yu; Larkin, Christopher J.; McGivney, James B.; Dovichi, Norman J.

    2016-01-01

    We have evaluated capillary zone electrophoresis-electrospray ionization-tandem mass spectrometry (CZE-ESI-MS/MS) for detection of trace amounts of host cell protein impurities in recombinant therapeutics. Compared to previously published procedures, we have optimized the buffer pH used in the formation of a pH junction to increase injection volume. We also prepared a five-point calibration curve by spiking twelve standard proteins into a solution of a human monoclonal antibody. A custom CZE-MS/MS system was used to analyze the tryptic digest of this mixture without depletion of the antibody. CZE generated a ~70 min separation window (~90 min total analysis duration) and ~300 peak capacity. We also analyzed the sample using ultra-performance liquid chromatography (UPLC)-MS/MS. CZE-MS/MS generated ~five times higher base peak intensity and more peptide identifications for low-level spiked proteins. Both methods detected all proteins spiked at the ~100 ppm level with respect to the antibody. PMID:26530276

  19. Comparative Glycoprofiling of HIV gp120 Immunogens by Capillary Electrophoresis and MALDI Mass Spectrometry

    Science.gov (United States)

    Guttman, Miklós; Váradi, Csaba; Lee, Kelly K.; Guttman, András

    2015-01-01

    The Human Immunodeficiency Virus (HIV) envelope glycoprotein (Env) is the primary antigenic feature on the surface of the virus and is of key importance in HIV vaccinology. Vaccine trials with the gp120 subunit of Env are ongoing with the recent RV144 trial showing moderate efficacy. gp120 is densely covered with N-linked glycans that are thought to help evade the host's humoral immune response. To assess how the global glycosylation patterns vary between gp120 constructs, the glycan profiles of several gp120s were examined by capillary electrophoresis with laser induced fluorescence detection and MALDI-MS. The glycosylation profiles were found to be similar for chronic vs. transmitter/founder isolates and only varied moderately between gp120s from different clades. This study revealed that the addition of specific tags, such as the gD tag used in the RV144 trial, had significant effects on the overall glycosylation patterns. Such effects are likely to influence the immunogenicity of various Env immunogens and should be considered for future vaccine strategies, emphasizing the importance of the glycosylation analysis approach described in this paper. PMID:25809283

  20. Simultaneous separation and determination of four uncaria alkaloids by capillary electrophoresis using dual cyclodextrin system.

    Science.gov (United States)

    Li, Lou; Xu, Liying; Chen, Meng; Zhang, Guangbin; Zhang, Hongfen; Chen, Anjia

    2017-07-15

    The purpose of this study was to develop a simple, quick and precise capillary zone electrophoresis method (CZE) for the separation and determination of uncaria alkaloids using dual cyclodextrins as additives for the separation. The four analytes were baseline separated within 15min at the applied voltage of 15kV with a running buffer (pH 5.7) consisting of 40.0mM phosphate buffer, 161.7mM 2-hydroxypropyl-β-cyclodextrin (HP-β-CD) and 2.21mM mono-(6-ethylenediamine-6-deoxy)-β-cyclodextrin (ED-β-CD). Under the optimum conditions, a good linearity was achieved with correlation coefficients from 0.9989 to 0.9992. The detection limits and the quantitation limits ranged from 0.63 to 0.98μg/mL and from 2.08 to 3.28μg/mL, respectively. Excellent accuracy and precision were obtained. Recoveries of the analytes varied from 97.1 to 103.2%. This method was suitable for the quantitative determination of these alkaloids in the stem with hook of Uncaria rhynchophylla and the formulations of Uncaria rhynchophylla. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Comparison of transferrin isoform analysis by capillary electrophoresis and HPLC for screening congenital disorders of glycosylation.

    Science.gov (United States)

    Dave, Mihika B; Dherai, Alpa J; Udani, Vrajesh P; Hegde, Anaita U; Desai, Neelu A; Ashavaid, Tester F

    2018-01-01

    Transferrin, a major glycoprotein has different isoforms depending on the number of sialic acid residues present on its oligosaccharide chain. Genetic variants of transferrin as well as the primary (CDG) & secondary glycosylation defects lead to an altered transferrin pattern. Isoform analysis methods are based on charge/mass variations. We aimed to compare the performance of commercially available capillary electrophoresis CDT kit for diagnosing congenital disorders of glycosylation with our in-house optimized HPLC method for transferrin isoform analysis. The isoform pattern of 30 healthy controls & 50 CDG-suspected patients was determined by CE using a Carbohydrate-Deficient Transferrin kit. The results were compared with in-house HPLC-based assay for transferrin isoforms. Transferrin isoform pattern for healthy individuals showed a predominant tetrasialo transferrin fraction followed by pentasialo, trisialo, and disialotransferrin. Two of 50 CDG-suspected patients showed the presence of asialylated isoforms. The results were comparable with isoform pattern obtained by HPLC. The commercial controls showed a <20% CV for each isoform. Bland Altman plot showed the difference plot to be within +1.96 with no systemic bias in the test results by HPLC & CE. The CE method is rapid, reproducible and comparable with HPLC and can be used for screening Glycosylation defects. © 2017 Wiley Periodicals, Inc.

  2. Metabolomics, peptidomics and proteomics applications of capillary electrophoresis-mass spectrometry in Foodomics: A review

    Energy Technology Data Exchange (ETDEWEB)

    Ibáñez, Clara; Simó, Carolina; García-Cañas, Virginia; Cifuentes, Alejandro, E-mail: a.cifuentes@csic.es; Castro-Puyana, María

    2013-11-13

    Graphical abstract: -- Highlights: •Foodomics allows studying food and nutrition through the application of advanced omics approaches. •CE-MS plays a crucial role as analytical platform to carry out omics studies. •CE-MS applications for food metabolomics, proteomics and peptidomics are presented. -- Abstract: In the current post-genomic era, Foodomics has been defined as a discipline that studies food and nutrition through the application of advanced omics approaches. Foodomics involves the use of genomics, transcriptomics, epigenetics, proteomics, peptidomics, and/or metabolomics to investigate food quality, safety, traceability and bioactivity. In this context, capillary electrophoresis-mass spectrometry (CE-MS) has been applied mainly in food proteomics, peptidomics and metabolomics. The aim of this review work is to present an overview of the most recent developments and applications of CE-MS as analytical platform for Foodomics, covering the relevant works published from 2008 to 2012. The review provides also information about the integration of several omics approaches in the new Foodomics field.

  3. Contribution of capillary electrophoresis to an integrated vision of humic substances size and charge characterizations

    International Nuclear Information System (INIS)

    D'Orlye, Fanny; Reiller, Pascal E.

    2014-01-01

    The physicochemical properties of three different humic substances (HS) are probed using capillary zone electrophoresis in alkaline carbonate buffers, pH 10. Special attention is drawn to the impact of the electrolyte ionic strength and counter-ion nature, chosen within the alkali-metal series, on HS electrophoretic mobility. Taylor-Aris dispersion analysis provides insights into the hydrodynamic radius (R-H) distributions of HS. The smallest characterized entities are of nano-metric dimensions, showing neither ionic strength- nor alkali-metal-induced aggregation. These results are compared with the entities evidenced in dynamic light scattering measurements, the size of which is two order of magnitude higher, ca. 100 nm. The extended Onsager model provides a reasonable description of measured electrophoretic mobilities in the ionic strength range 1-50 mM, thus allowing the estimation of limiting mobilities and ionic charge numbers for the different HS samples. An unexpected HS electrophoretic mobility increase (in absolute value) is observed in the order Li + ≤ Na + ≤ K + ≤ Cs + and discussed either in terms of retarding forces or in terms of ion-ion interactions. (authors)

  4. Capillary electrophoresis single-strand conformation polymorphism for the monitoring of gastrointestinal microbiota of chicken flocks.

    Science.gov (United States)

    Pissavin, C; Burel, C; Gabriel, I; Beven, V; Mallet, S; Maurice, R; Queguiner, M; Lessire, M; Fravalo, P

    2012-09-01

    The objective of the present study was to evaluate the capillary electrophoresis single-strand conformation polymorphism (CE-SSCP) to characterize poultry gut microbiota and the ability of this molecular method to detect modifications related to rearing conditions to be used as an epidemiological tool. The V3 region of the 16S rRNA gene was selected as the PCR target. Our results showed that this method provides reproducible data. The microbiota analysis of individuals showed that variability between individual fingerprints was higher for ileum and cloaca than for ceca. However, pooling the samples decreased this variability. To estimate the variability within and between farms, we compared molecular gut patterns of animals from the same hatchery reared under similar conditions and fed the same diet in 2 separate farms. Total aerobic bacteria, coliforms, and lactic acid bacteria were enumerated using conventional bacteriological methods. A significant difference was observed for coliforms present in the ceca and the cloaca depending on the farm. Ileal contents fingerprints were more closely related to those of cloacal contents than to those of ceca contents. When comparing samples from the 2 farms, a specific microbiota was highlighted for each farm. For each gut compartment, the microbiota fingerprints were joined in clusters according to the farm. Thus, this rapid and potentially high-throughput method to obtain gut flora fingerprints is sensitive enough to detect a "farm effect" on the balance of poultry gut microbiota despite the birds being fed the same regimens and reared under similar conditions.

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

    Directory of Open Access Journals (Sweden)

    Young Chon Choi

    2006-06-01

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

  6. Capillary electrophoresis - inductively coupled plasma mass spectrometry (CE-ICPMS) coupling to assess pentavalent actinides thermodynamic constants

    International Nuclear Information System (INIS)

    Topin, S.; Baglan, N.; Aupiais, J.

    2009-01-01

    Full text: Aiming to investigate plutonium speciation at trace levels, we coupled capillary electrophoresis, a high resolution separation technique with inductively coupled plasma mass spectrometry, a detector with high sensitivity for plutonium. The research work performed to optimize the coupling is discussed based on the following criteria: the migration time, the resolution and the detection limit. The capabilities of the analytical tool are demonstrated by determining thermodynamic constants for pentavalent plutonium, and neptunium as a reference, in the presence of inorganic ligands. (author)

  7. Simultaneous determination of anthraquinones, their 8-beta-D-glucosides, and sennosides of Rhei Rhizoma by capillary electrophoresis.

    Science.gov (United States)

    Koyama, Junko; Morita, Izumi; Fujiyoshi, Hirotaka; Kobayashi, Norihiro

    2005-05-01

    The simultaneous separation and determination of major anthraquinones (emodin, chrysophanol, rhein and their glucosides, aloe-emodin, sennoside A, and sennoside B) of Rhei Rhizoma were achieved by cyclodextrin modified capillary zone electrophoresis. The running electrolyte used in this method was 0.005 M alpha-cyclodextrin in 0.03 M borate buffer (pH 10.0) containing 20% acetonitrile, with an applied voltage of 20 kV.

  8. Determination of anions with an on-line capillary electrophoresis method; Anionien on-line maeaeritys kapillaarielektroforeesilla - MPKT 10

    Energy Technology Data Exchange (ETDEWEB)

    Siren, H; Saerme, T; Kotiaho, T; Hiissa, T; Savolahti, P; Komppa, V [VTT Chemical Technology, Espoo (Finland)

    1999-12-31

    The aim of the study was to set-up an on-line capillary electrophoresis method for determination of anions in process waters of pulp and paper industry with exporting the results to the process control system of the mill. The quantification is important, since it will give information about the possible causes of precipitation. In recent years, the capillary electrophoresis (CE) due to its high separation efficiency has been shown as a method to take into consideration when analyzing chemical species ranging from small inorganic anions to different macromolecules. Many compounds are not easily detected in their native state, why analysis methods must be developed to improve their detection. Especially, small inorganic and organic anions which do not have chromophores are not sensitive enough for direct-UV detection. In such analyses the anions are mostly detected with indirect-UV technique. Capillary electrophoresis instruments are used to analyze samples in off-line, which seldom represent the situation in process. Therefore, on-line instrument technology with autoanalyzing settings will be needed in quality control. The development of a fully automatic capillary electrophoresis system is underway in co-operation with KCL (The Finnish Pulp and Paper Research Institute). In our research, we have first concentrated on the determination of sulphate in waters of paper industry. The method used for detection of sulphate is based on indirect-UV detection with CE, where the background electrolyte (BGE) is an absorbing mixture of secondary amines. The whole procedure for quantification of sulphate is performed within 15 minutes, after which a new sample is analyzed automatically. The only sample pretreatment is filtration, which is necessary before analysis. The concentrations of sulphate in process waters tested were between 300 and 800 ppm. Our tests show that a simultaneous determination of chloride, sulphate, nitrate, nitrite, sulphite, carbonate and oxalate is also

  9. Determination of anions with an on-line capillary electrophoresis method; Anionien on-line maeaeritys kapillaarielektroforeesilla - MPKT 10

    Energy Technology Data Exchange (ETDEWEB)

    Siren, H.; Saerme, T.; Kotiaho, T.; Hiissa, T.; Savolahti, P.; Komppa, V. [VTT Chemical Technology, Espoo (Finland)

    1998-12-31

    The aim of the study was to set-up an on-line capillary electrophoresis method for determination of anions in process waters of pulp and paper industry with exporting the results to the process control system of the mill. The quantification is important, since it will give information about the possible causes of precipitation. In recent years, the capillary electrophoresis (CE) due to its high separation efficiency has been shown as a method to take into consideration when analyzing chemical species ranging from small inorganic anions to different macromolecules. Many compounds are not easily detected in their native state, why analysis methods must be developed to improve their detection. Especially, small inorganic and organic anions which do not have chromophores are not sensitive enough for direct-UV detection. In such analyses the anions are mostly detected with indirect-UV technique. Capillary electrophoresis instruments are used to analyze samples in off-line, which seldom represent the situation in process. Therefore, on-line instrument technology with autoanalyzing settings will be needed in quality control. The development of a fully automatic capillary electrophoresis system is underway in co-operation with KCL (The Finnish Pulp and Paper Research Institute). In our research, we have first concentrated on the determination of sulphate in waters of paper industry. The method used for detection of sulphate is based on indirect-UV detection with CE, where the background electrolyte (BGE) is an absorbing mixture of secondary amines. The whole procedure for quantification of sulphate is performed within 15 minutes, after which a new sample is analyzed automatically. The only sample pretreatment is filtration, which is necessary before analysis. The concentrations of sulphate in process waters tested were between 300 and 800 ppm. Our tests show that a simultaneous determination of chloride, sulphate, nitrate, nitrite, sulphite, carbonate and oxalate is also

  10. Affinity capillary electrophoresis and density functional theory study of noncovalent interactions of cyclic peptide [Gly(6)]-antamanide with small cations

    Czech Academy of Sciences Publication Activity Database

    Pangavhane, Sachin; Böhm, S.; Makrlík, E.; Ruzza, P.; Kašička, Václav

    2017-01-01

    Roč. 38, č. 16 (2017), s. 2025-2033 ISSN 0173-0835 R&D Projects: GA ČR(CZ) GA15-01948S Institutional support: RVO:61388963 Keywords : affinity capillary electrophoresis * cesium complex * density functional theory * [Gly(6)]-antamanide * rubidium complex * stability constants Subject RIV: CB - Analytical Chemistry, Separation OBOR OECD: Analytical chemistry Impact factor: 2.744, year: 2016

  11. Optimization of the Nonaqueous Capillary Electrophoresis Separation of Metal Ions Using Mixture Design and Response Surface Methods

    OpenAIRE

    DEMİR, Cevdet; YÜCEL, Yasin

    2014-01-01

    Mixture experimental design was used to enhance the separation selectivity of metal ions in nonaqueous capillary electrophoresis. The separation of cations (Ag, Fe, Cr, Mn, Cd, Co, Pb, Ni, Zn and Cu) was achieved using imidazole as UV co-ion for indirect detection. Acetic acid was chosen as an electrolyte because its cathodic electroosmotic flow permits faster separation. The composition of organic solvents is important to achieve the best separation of all metal ions. Simplex latt...

  12. Review: Authentication and traceability of foods from animal origin by polymerase chain reaction-based capillary electrophoresis.

    Science.gov (United States)

    Rodríguez-Ramírez, Roberto; González-Córdova, Aarón F; Vallejo-Cordoba, Belinda

    2011-01-31

    This work presents an overview of the applicability of PCR-based capillary electrophoresis (CE) in food authentication and traceability of foods from animal origin. Analytical approaches for authenticating and tracing meat and meat products and fish and seafood products are discussed. Particular emphasis will be given to the usefulness of genotyping in food tracing by using CE-based genetic analyzers. Copyright © 2010 Elsevier B.V. All rights reserved.

  13. Determination of acid-base dissociation constants of very weak zwitterionic heterocyclic bases by capillary zone electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Ehala, Sille; Grishina, Anastasia; Sheshenev, Andrey; Lyapkalo, Ilya; Kašička, Václav

    2010-01-01

    Roč. 1217, - (2010), s. 8048-8053 ISSN 0021-9673 R&D Projects: GA ČR(CZ) GA203/08/1428; GA ČR(CZ) GA203/09/0675 Institutional research plan: CEZ:AV0Z40550506 Keywords : acidity constant * capillary zone electrophoresis * zwitterionic heterocyclic bases Subject RIV: CC - Organic Chemistry Impact factor: 4.194, year: 2010

  14. Determination of methicillin-resistant and methicillin-susceptible Staphylococcus aureus bacteria in blood by capillary zone electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Horká, Marie; Tesařová, Marie; Karásek, Pavel; Růžička, F.; Holá, V.; Sittová, M.; Roth, Michal

    2015-01-01

    Roč. 868, APR (2015), s. 67-72 ISSN 0003-2670 R&D Projects: GA ČR(CZ) GAP106/12/0522; GA MV VG20102015023 Institutional support: RVO:68081715 Keywords : capillary zone electrophoresis * Staphylococcus aureus * human whole blood Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 4.712, year: 2015 http://hdl.handle.net/11104/0245801

  15. Estimation of apparent binding constant of complexes of selected acyclic nucleoside phosphonates with beta-cyclodextrin by affinity capillary electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Šolínová, Veronika; Mikysková, Hana; Kaiser, Martin Maxmilian; Janeba, Zlatko; Holý, Antonín; Kašička, Václav

    2016-01-01

    Roč. 37, č. 2 (2016), s. 239-247 ISSN 0173-0835 R&D Projects: GA ČR(CZ) GA13-17224S; GA ČR(CZ) GA15-01948S Institutional support: RVO:61388963 Keywords : acyclic nucleoside phosphonates * affinity capillary electrophoresis * binding constant * nucleotide analogs * beta-cyclodextrin Subject RIV: CB - Analytical Chemistry , Separation Impact factor: 2.744, year: 2016

  16. Electrophoresis gel image processing and analysis using the KODAK 1D software.

    Science.gov (United States)

    Pizzonia, J

    2001-06-01

    The present article reports on the performance of the KODAK 1D Image Analysis Software for the acquisition of information from electrophoresis experiments and highlights the utility of several mathematical functions for subsequent image processing, analysis, and presentation. Digital images of Coomassie-stained polyacrylamide protein gels containing molecular weight standards and ethidium bromide stained agarose gels containing DNA mass standards are acquired using the KODAK Electrophoresis Documentation and Analysis System 290 (EDAS 290). The KODAK 1D software is used to optimize lane and band identification using features such as isomolecular weight lines. Mathematical functions for mass standard representation are presented, and two methods for estimation of unknown band mass are compared. Given the progressive transition of electrophoresis data acquisition and daily reporting in peer-reviewed journals to digital formats ranging from 8-bit systems such as EDAS 290 to more expensive 16-bit systems, the utility of algorithms such as Gaussian modeling, which can correct geometric aberrations such as clipping due to signal saturation common at lower bit depth levels, is discussed. Finally, image-processing tools that can facilitate image preparation for presentation are demonstrated.

  17. Streamlined sign-out of capillary protein electrophoresis using middleware and an open-source macro application.

    Science.gov (United States)

    Mathur, Gagan; Haugen, Thomas H; Davis, Scott L; Krasowski, Matthew D

    2014-01-01

    Interfacing of clinical laboratory instruments with the laboratory information system (LIS) via "middleware" software is increasingly common. Our clinical laboratory implemented capillary electrophoresis using a Sebia(®) Capillarys-2™ (Norcross, GA, USA) instrument for serum and urine protein electrophoresis. Using Data Innovations Instrument Manager, an interface was established with the LIS (Cerner) that allowed for bi-directional transmission of numeric data. However, the text of the interpretive pathology report was not properly transferred. To reduce manual effort and possibility for error in text data transfer, we developed scripts in AutoHotkey, a free, open-source macro-creation and automation software utility. Scripts were written to create macros that automated mouse and key strokes. The scripts retrieve the specimen accession number, capture user input text, and insert the text interpretation in the correct patient record in the desired format. The scripts accurately and precisely transfer narrative interpretation into the LIS. Combined with bar-code reading by the electrophoresis instrument, the scripts transfer data efficiently to the correct patient record. In addition, the AutoHotKey script automated repetitive key strokes required for manual entry into the LIS, making protein electrophoresis sign-out easier to learn and faster to use by the pathology residents. Scripts allow for either preliminary verification by residents or final sign-out by the attending pathologist. Using the open-source AutoHotKey software, we successfully improved the transfer of text data between capillary electrophoresis software and the LIS. The use of open-source software tools should not be overlooked as tools to improve interfacing of laboratory instruments.

  18. Streamlined sign-out of capillary protein electrophoresis using middleware and an open-source macro application

    Directory of Open Access Journals (Sweden)

    Gagan Mathur

    2014-01-01

    Full Text Available Background: Interfacing of clinical laboratory instruments with the laboratory information system (LIS via "middleware" software is increasingly common. Our clinical laboratory implemented capillary electrophoresis using a Sebia; Capillarys-2™ (Norcross, GA, USA instrument for serum and urine protein electrophoresis. Using Data Innovations Instrument Manager, an interface was established with the LIS (Cerner that allowed for bi-directional transmission of numeric data. However, the text of the interpretive pathology report was not properly transferred. To reduce manual effort and possibility for error in text data transfer, we developed scripts in AutoHotkey, a free, open-source macro-creation and automation software utility. Materials and Methods: Scripts were written to create macros that automated mouse and key strokes. The scripts retrieve the specimen accession number, capture user input text, and insert the text interpretation in the correct patient record in the desired format. Results: The scripts accurately and precisely transfer narrative interpretation into the LIS. Combined with bar-code reading by the electrophoresis instrument, the scripts transfer data efficiently to the correct patient record. In addition, the AutoHotKey script automated repetitive key strokes required for manual entry into the LIS, making protein electrophoresis sign-out easier to learn and faster to use by the pathology residents. Scripts allow for either preliminary verification by residents or final sign-out by the attending pathologist. Conclusions: Using the open-source AutoHotKey software, we successfully improved the transfer of text data between capillary electrophoresis software and the LIS. The use of open-source software tools should not be overlooked as tools to improve interfacing of laboratory instruments.

  19. 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.

  20. Native gel electrophoresis of human telomerase distinguishes active complexes with or without dyskerin

    Science.gov (United States)

    Gardano, Laura; Holland, Linda; Oulton, Rena; Le Bihan, Thierry; Harrington, Lea

    2012-01-01

    Telomeres, the ends of linear chromosomes, safeguard against genome instability. The enzyme responsible for extension of the telomere 3′ terminus is the ribonucleoprotein telomerase. Whereas telomerase activity can be reconstituted in vitro with only the telomerase RNA (hTR) and telomerase reverse transcriptase (TERT), additional components are required in vivo for enzyme assembly, stability and telomere extension activity. One such associated protein, dyskerin, promotes hTR stability in vivo and is the only component to co-purify with active, endogenous human telomerase. We used oligonucleotide-based affinity purification of hTR followed by native gel electrophoresis and in-gel telomerase activity detection to query the composition of telomerase at different purification stringencies. At low salt concentrations (0.1 M NaCl), affinity-purified telomerase was ‘supershifted’ with an anti-dyskerin antibody, however the association with dyskerin was lost after purification at 0.6 M NaCl, despite the retention of telomerase activity and a comparable yield of hTR. The interaction of purified hTR and dyskerin in vitro displayed a similar salt-sensitive interaction. These results demonstrate that endogenous human telomerase, once assembled and active, does not require dyskerin for catalytic activity. Native gel electrophoresis may prove useful in the characterization of telomerase complexes under various physiological conditions. PMID:22187156

  1. Evaluation of iron and selenium losses in metalloproteins separated by gel electrophoresis by ICPMS

    International Nuclear Information System (INIS)

    Gherghel, I.; Fernandez, M.L.; Fernandez, B.; Pereiro, R.; Sanz-Medel, A.

    2009-01-01

    Full text: Metallomics addresses the study of metabolism, transport, and metal-protein interactions aiming to obtain relevant information of physiological and pathological alterations in living organisms. Gel electrophoresis is widely employed in proteomics and its use is actually extending to metallomics. Unfortunately, analysis of proteins by molecular techniques does not offer quantitative information. So, a good alternative could be their determination via the quantification of (semi)metal bound to the protein by ICPMS. In this work, we will show a detailed study of possible losses of protein and/or metal in Fe-bound and selenium proteins (transferrin and glutathione peroxidase, respectively) to evaluate the behaviour of the protein-metal interactions during the electrophoresis process. (author)

  2. Alkaline gel electrophoresis assay to detect DNA strand breaks and repair mechanisms in Escherichia coli

    Directory of Open Access Journals (Sweden)

    José Carlos Pelielo de Mattos

    2008-12-01

    Full Text Available Reactive oxygen species (ROS can induce lesions in different cellular targets, including DNA. Stannous chloride (SnCl2 is a ROS generator, leading to lethality in Escherichia coli (E. coli, with the base excision repair (BER mechanism playing a role in this process. Many techniques have been developed to detect genotoxicity, as comet assay, in eukaryotic cells, and plasmid DNA agarose gel electrophoresis. In this study, an adaptation of the alkaline gel electrophoresis method was carried out to ascertain the induction of strand breaks by SnCl2 in bacterial DNA, from E. coli BER mutants, and its repair pathway. Results obtained show that SnCl2 was able to induce DNA strand breaks in all strains tested. Moreover, endonuclease IV and exonuclease III play a role in DNA repair. On the whole, data has shown that the alkaline gel electrophoresis assay could be used both for studying DNA strand breaks induction and for associated repair mechanisms.Espécies reativas de oxigênio (ERO podem induzir lesões em diferentes alvos celulares, incluindo o DNA. O cloreto estanoso (SnCl2 é um gerador de ERO que induz letalidade em E. coli, sendo o reparo por excisão de bases (BER um mecanismo importante neste processo. Técnicas como o ensaio cometa (em eucariotos e a eletroforese de DNA plasmidial em gel de agarose têm sido utilizadas para detectar genotoxicidade. No presente estudo, uma adaptação do método de eletroforese em gel alcalino de agarose foi usada para verificar a indução de quebras, pelo SnCl2, no DNA de E. coli, bem como a participação de enzimas do BER na restauração das lesões. Os resultados mostraram que o SnCl2 induziu quebras no DNA de todas as cepas testadas. Além disso, endonuclease IV e exonuclease III estão envolvidas na reparação dos danos. Em resumo, os dados obtidos indicam que a metodologia de eletroforese em gel alcalino de agarose pode ser empregada tanto para o estudo de quebras no DNA, quanto para avaliação dos

  3. pI-Control in comparative fluorescence gel electrophoresis (CoFGE) using amphoteric azo dyes

    Czech Academy of Sciences Publication Activity Database

    Hanneken, M.; Šlais, Karel; König, S.

    2015-01-01

    Roč. 8, SEP (2015), s. 36-39 ISSN 2212-9685 Institutional support: RVO:68081715 Keywords : comparative fluorescence gel * electrophoresis * protein grid * azo dyes Subject RIV: CB - Analytical Chemistry, Separation http://ac.els-cdn.com/S2212968515000094/1-s2.0-S2212968515000094-main.pdf?_tid=7c92fa40-56e6-11e5-b36a-00000aab0f01&acdnat=1441798543_19612c0d7466780944bc4ae22173da92

  4. Analysis of the surface membrane of iodinated leukemic cells by SDS-polyacrylamide gel electrophoresis

    International Nuclear Information System (INIS)

    Ishitani, Kunihiko; Ikeda, Akira; Tamura, Minoru; Takeuchi, Hidekazu; Ihara, Koji

    1980-01-01

    Surface proteins of human leukemic cells were labeled selectively by lactoperoxydase catalysed-iodination and examined by SDS-polyacrylamide gel electrophoresis. The electrophoretic pattern of the surface membranes of cells from a patients with chronic mylogeneous leukemia in blast crisis was of B cell type and showed Ia like antigen. Leukemic cells from a patient with hairly cell leukemia also expressed the pattern of B cell type when tested by this method the technique of iodinating cell surface with lactoperoxidase is useful in characterization of leukemia cells for diagnosis and monitoring of clinical course. (author)

  5. The use of the 2-aminobenzoic acid tag for oligosaccharide gel electrophoresis.

    Science.gov (United States)

    Huang, Z; Prickett, T; Potts, M; Helm, R F

    2000-08-18

    Gel electrophoresis of fluorophore labeled saccharides provides a rapid and reliable method to screen enzymatic and/or chemical treatments of polysaccharides and glycoconjugates, as well as a sensitive and efficient microscale method to separate and purify oligosaccharides for further analysis. A simple and inexpensive method of derivatization and analysis using 2-aminobenzoic acid (anthranilic acid, AA) is described and applied to the extracellular polysaccharide released by the desiccation tolerant cyanobacterium Nostoc commune DRH-1. The results of these analyses suggest a possible protective functionality of two pendent groups, as well as a potential relationship between these groups and the desiccation tolerance of the organism.

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

    Directory of Open Access Journals (Sweden)

    Perez-Chabela, M. Lourdes

    2015-12-01

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

  7. Direct detection of rutin-degrading isozymes with polyacrylamide gel electrophoresis.

    Science.gov (United States)

    Li, Yuping; Deng, Dandan; Zhang, Xuebin; Zhang, Haina; Wang, Cong; Chen, Peng

    2013-12-15

    Rutin-degrading enzymes (RDEs) specifically hydrolyze the glycosidic linkages of rutin, producing quercetin and rutinose. Here we report a reliable and sensitive polyacrylamide gel electrophoresis and staining method for the detection of RDE isozymes, which is based on the aqueous solubility difference between rutin and quercetin, as well as the ultraviolet absorbance of quercetin. With this novel method, we achieved a detection limit of 12 ng with 107 U of RDE activity, enabling us to detect at least five RDE isozymes in tartary buckwheat seeds. Copyright © 2013 Elsevier Inc. All rights reserved.

  8. Study on the non-target effect of ionizing radiation using single cell gel electrophoresis

    International Nuclear Information System (INIS)

    Wang Yan; Li Deguan; Liu Jianfeng; Chu Liping; Liu Qiang

    2008-01-01

    Objective: To assess the non-target effect of ionizing radiation by single cell gel electrophoresis (SCGE). Methods: Cross incubated the irradiated( 137 Cs; 2Gy) or non-irradiated lymphocytes of human peripheral blood in the irradiated or non-irradiated plasma respectively, then, assess the DNA damage of lymphocytes using SCGE analysis. Results: The lymphocytes incubated in the irradiated plasma presented more obvious DNA damage than the incubated in the non-irradiated plasma dose (P<0.05). Conclusion: The non-target effect of ionizing radiation can be assessed by SCGE, and the results confirm that cytokines may play a great role in it. (authors)

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

    International Nuclear Information System (INIS)

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

    2010-01-01

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

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

    DEFF Research Database (Denmark)

    Christiansen, L; Ged, C; Hombrados, I

    1999-01-01

    to exon skipping, and a 2-bp deletion (415-416delTA) resulting in a frameshift and the introduction of a premature stop codon. Heterologous expression and enzymatic studies of the mutant proteins demonstrate that the three mutations leading to shortening or truncation of the UROD protein have no residual......, confirming the heterogeneity of the underlying genetic defects of these diseases. We have established a denaturing gradient gel electrophoresis (DGGE) assay for mutation detection in the UROD gene, enabling the simultaneous screening for known and unknown mutations. The established assay has proved able...

  11. The limitations of pulsed-field gel electrophoresis for analysis of Yersinia enterocolitica isolates.

    Science.gov (United States)

    Gilpin, B J; Robson, B; Lin, S; Hudson, J A; Weaver, L; Dufour, M; Strydom, H

    2014-09-01

    This study describes the analysis of 432 isolates of Yersinia enterocolitica by pulsed-field gel electrophoresis (PFGE). PFGE had a high level of discrimination with biotype 1A isolates (Simpson's Diversity Index 0.997), but with the clinically important biotypes 2, 3 and 4, the discriminatory ability of PFGE was so low as to severely limit its usefulness (DI enterocolitica biotypes 2, 3 and 4, and inferences based on finding indistinguishable PFGE profiles among cases or between cases and sources need to be substantiated using alternative typing tools, or strong epidemiological evidence. © 2013 Blackwell Verlag GmbH.

  12. Using nonequilibrium capillary electrophoresis of equilibrium mixtures (NECEEM) for simultaneous determination of concentration and equilibrium constant.

    Science.gov (United States)

    Kanoatov, Mirzo; Galievsky, Victor A; Krylova, Svetlana M; Cherney, Leonid T; Jankowski, Hanna K; Krylov, Sergey N

    2015-03-03

    Nonequilibrium capillary electrophoresis of equilibrium mixtures (NECEEM) is a versatile tool for studying affinity binding. Here we describe a NECEEM-based approach for simultaneous determination of both the equilibrium constant, K(d), and the unknown concentration of a binder that we call a target, T. In essence, NECEEM is used to measure the unbound equilibrium fraction, R, for the binder with a known concentration that we call a ligand, L. The first set of experiments is performed at varying concentrations of T, prepared by serial dilution of the stock solution, but at a constant concentration of L, which is as low as its reliable quantitation allows. The value of R is plotted as a function of the dilution coefficient, and dilution corresponding to R = 0.5 is determined. This dilution of T is used in the second set of experiments in which the concentration of T is fixed but the concentration of L is varied. The experimental dependence of R on the concentration of L is fitted with a function describing their theoretical dependence. Both K(d) and the concentration of T are used as fitting parameters, and their sought values are determined as the ones that generate the best fit. We have fully validated this approach in silico by using computer-simulated NECEEM electropherograms and then applied it to experimental determination of the unknown concentration of MutS protein and K(d) of its interactions with a DNA aptamer. The general approach described here is applicable not only to NECEEM but also to any other method that can determine a fraction of unbound molecules at equilibrium.

  13. Bioanalysis of tobramycin for therapeutic drug monitoring by solid-phase extraction and capillary zone electrophoresis.

    Science.gov (United States)

    Fonge, Humphrey; Kaale, Eliangiringa; Govaerts, Cindy; Desmet, Koenraad; Van Schepdael, Ann; Hoogmartens, Jos

    2004-10-25

    A method based on solid-phase extraction (SPE) and capillary zone electrophoresis (CZE) for the analysis of tobramycin in human serum is presented. An off-line SPE employing a carboxypropyl bonded phase (CBA) cartridge was used for the extraction of tobramycin from human serum. Adsorbed tobramycin was eluted from the CBA cartridge using a mixture of NH(3) (25%, w/v)-methanol (30:70, v/v). After evaporation, the analyte was reconstituted and derivatized with o-phthaldialdehyde (OPA)/3-mercaptopropionic acid (MPA). The resulting tobramycin-OPA/MPA derivative was purified, and then identified by mass spectrometry. The tobramycin-OPA/MPA derivative was then analysed by CZE with a background electrolyte (BGE) comprising of 30 mM sodium tetraborate pH 10.0-acetonitrile (ACN) (80:20, v/v) with ultraviolet detection at 230 nm. A linear response was observed in the range of 0.3-30 microg/ml with r(2) = 0.992. The sensitivity of the method was determined by its limit of quantitation (LOQ) and limit of detection (LOD) of 0.3 microg/ml and 0.1 microg/ml, respectively. SPE recovery ranged from 68 to 79% at the trough levels to 98% at the peak levels found in serum. Furosemide has been added as internal standard (IS) to improve precision. For the therapeutic range of tobramycin in serum (2-10 microg/ml) the relative standard deviation (R.S.D.) was less than 11% for the entire SPE/CE process. The method demonstrated excellent selectivity as shown by the lack of interference from a total of 20 drugs investigated. The method was then used in therapeutic drug monitoring of patients receiving the drug.

  14. Simultaneous determination of psychotropic drugs in human urine by capillary electrophoresis with electrochemiluminescence detection

    Energy Technology Data Exchange (ETDEWEB)

    Li Jianguo [Key Laboratory of Analytical Chemistry for Life Science (Education Ministry of China), School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210093 (China); School of Chemistry and Chemical Engineering, Suzhou University, Suzhou 215006 (China); Zhao Fengjuan [Key Laboratory of Analytical Chemistry for Life Science (Education Ministry of China), School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210093 (China); Ju Huangxian [Key Laboratory of Analytical Chemistry for Life Science (Education Ministry of China), School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210093 (China)]. E-mail: hxju@nju.edu.cn

    2006-08-04

    Amitriptyline, doxepin and chlorpromazine are often used as psychotropic drugs in treatment of the various mental diseases, and are also partly excreted by kidney. This work developed a simple, selective and sensitive method for their simultaneous monitoring in human urine using capillary electrophoresis coupled with electrochemiluminescence (ECL) detection based on end-column ECL reaction of tris-(2,2'-bipyridyl)ruthenium(II) with aliphatic tertiary amino moieties. Acetone was used as an additive to the running buffer to obtain their absolute separation. Under optimized conditions the proposed method displayed a linear range from 5.0 to 800 ng mL{sup -1} for the three drugs with the correlation coefficients more than 0.995 (n = 8). Their limits of detection were 0.8 ng mL{sup -1} (3.6 fg), 1.0 ng mL{sup -1} (4.5 fg) and 1.5 ng mL{sup -1} (6.8 fg) at a signal to noise ratio of 3, respectively. The relative standard deviations for five determinations of 20 ng mL{sup -1} amitriptyline, doxepin and chlorpromazine were 1.7%, 4.2% and 3.6%, respectively. For practical application an extract step with 90:10 heptane/ethyl acetate (v/v) was performed to eliminate the influence of ionic strength in sample. The recoveries of amitriptyline, doxepin and chlorpromazine at different levels in human urine were between 83% and 93%, which showed that the method was valuable in clinical and biochemical laboratories for monitoring amitriptyline, doxepin and chlorpromazine.

  15. Chiral discrimination of sibutramine enantiomers by capillary electrophoresis and proton nuclear magnetic resonance spectroscopy.

    Science.gov (United States)

    Lee, Yong-Jae; Choi, Seungho; Lee, Jinhoo; Nguyen, NgocVan Thi; Lee, Kyungran; Kang, Jong Seong; Mar, Woongchon; Kim, Kyeong Ho

    2012-03-01

    Capillary electrophoresis (CE) and proton nuclear magnetic resonance spectroscopy ((1)H-NMR) have been used to discriminate the enantiomers of sibutramine using cyclodextrin derivatives. Possible correlation between CE and (1)H-NMR was examined. Good correlation between the (1)H-NMR shift non-equivalence data for sibutramine and the degree of enantioseparation in CE was observed. In CE study, a method of enantiomeric separation and quantitation of sibutramine was developed using enantiomeric standards. The method was based on the use of 50 mM of phosphate buffer of pH 3.0 with 10 mM of methyl-beta-cyclodextrin (M-β-CD). 0.05% of LOD, 0.2% of LOQ for S-sibutramine enantiomer was achieved, and the method was validated and applied to the quantitative determination of sibutramine enantiomers in commercial drugs. On a 600 MHz (1)H-NMR analysis, enantiomer signal separation of sibutramine was obtained by fast diastereomeric interaction with a chiral selector M-β-CD. For chiral separation and quantification, N-methyl proton peaks (at 2.18 ppm) were selected because of its being singlet and simple for understanding of diastereomeric interaction. Effects of temperature and concentration of chiral selector on enantiomer signal separation were investigated. The optimum condition was 0.5 mg/mL of sibutramine and 10 mg/mL of M-β-CD at 10°C. Distinguishment of 0.5% of S-sibutramine in R-sibutramine was found to be possible by (1)H-NMR with M-β-CD as chiral selector. Host-guest interaction between sibutramine and M-β-CD was confirmed by (1)H-NMR studies and CE studies. A Structure of the inclusion complex was proposed considering (1)H-NMR and 2D ROESY studies.

  16. Speciation of the plutonium at trace levels by capillary electrophoresis-ICP-MS coupling

    International Nuclear Information System (INIS)

    Ambard, Ch.

    2007-01-01

    The CE-ICP-MS coupling allowed the development of new analytical methods for the study of plutonium (Pu) speciation at trace levels including complexation studies of this element by organic and inorganic ligands. First, a method, called dual detection, based on the simultaneous use of the UV-Visible spectrophotometer integrated in the capillary electrophoresis and the ICP-MS was developed and validated. It allows the unambiguous determination of electrophoretic mobilities for separated chemical species and gives a powerful tool for speciation studies. Then, the influence on Pu redox speciation of the buffer from the background electrolyte was evaluated. This study showed the implications of the electrolyte constituents' choice on Pu redox equilibrium in the sample. Furthermore, the CE-ICP-MS coupling was used for studying the Pu complexation at trace levels by some organic (NTA and DTPA) and inorganic ligands (carbonates). The behaviour of Pu valence +III, +IV and +VI was studied in the presence of buffer at near neutral pH. Different species of Pu were observed depending on the initial oxidation state of the plutonium. The study showed the potential of poly-amino-carboxylic acids, such as NTA and DTPA, for dissolving Pu precipitates, regardless its initial speciation. Finally, the carbonation of pentavalent neptunium, as an analogue of Pu(V), was achieved at very low concentration of Np (10 -8 mol.L -1 ). The formation of NpO 2 (CO 3 ) - at 25 C and 2,5*10 -2 mol.L -1 ionic strength was measured by CE-ICP-MS and found to consistent with literature data. (author)

  17. Speciation of the plutonium at trace levels by capillary electrophoresis-ICP-MS coupling

    International Nuclear Information System (INIS)

    Ambard, Ch.

    2007-01-01

    The CE-ICP-MS coupling allowed the development of new analytical methods for the study of plutonium speciation at trace levels including complexation studies of this element by organic and inorganic ligands. First, a method, called dual detection, based on the simultaneous use of the UV-Visible spectrophotometer integrated in the capillary electrophoresis and the ICPMS was developed and validated. It allows the unambiguous determination of electrophoretic mobilities for separated chemical species and gives a powerful tool for speciation studies. Then, the influence on plutonium redox speciation of the buffer from the background electrolyte was evaluated. This study showed the implications of the electrolyte constituents' choice on plutonium redox equilibrium in the sample. Furthermore, the CE-ICP-MS coupling was used for studying the plutonium complexation at trace levels by some organic (NTA and DTPA) and inorganic ligands (carbonates). The behaviour of plutonium valence +III, +IV and +VI was studied in the presence of buffer at near neutral pH. Different species of plutonium were observed depending on the initial oxidation state of the plutonium. This study showed the potential of poly-amino-carboxylic acids, such as NTA and DTPA, for dissolving plutonium precipitates, regardless its initial speciation. Finally, the carbonation of pentavalent neptunium, as an analogue of Pu(V), was achieved at very low concentration of Np (10 -8 mol.L -1 ). The formation constant of NpO 2 (CO 3 )- at 25 deg. C and 2,5 x 10 -2 mol.L -1 ionic strength was measured by CE-ICP-MS and found to be consistent with literature data. (author)

  18. Development of methods to measure hemoglobin adducts by gel electrophoresis - Preliminary results

    International Nuclear Information System (INIS)

    Sun, J.D.; McBride, S.M.

    1988-01-01

    Chemical adducts formed on blood hemoglobin may be a useful biomarker for assessing human exposures to these compounds. This paper reports preliminary results in the development of methods to measure such adducts that may be generally applicable for a wide variety of chemicals. Male F344/N rats were intraperitoneally injected with 14 C-BaP dissolved in corn oil. Twenty-four hours later, the rats were sacrificed. Blood samples were collected and globin was isolated. Globin protein was then cleaved into peptide fragments using cyanogen bromide and the fragments separated using 2-dimensional gel electrophoresis. The results showed that the adducted 14 C-globin fragments migrated to different areas of the gel than did unadducted fragments. Further research is being conducted to develop methods that will allow quantitation of separated adducted globin fragments from human blood samples without the use of a radiolabel. (author)

  19. Sodium Dodecyl Sulfate- Polyacrylamide Gel Electrophoresis (SDS- PAGE) of Irradiated Wheat Flour Proteins

    International Nuclear Information System (INIS)

    Souzan, R.M.

    1999-01-01

    Sodium dodecyl sulfate - polyacrylamide gel electrophoresis (SDS-PAGE) of wheat (Triticum aestivum L) flour have revealed 23 polypeptides of molecular weights between 170 and 11.57 KDa, High molecular weight glutenin subunits (LMW-GS) were distinguished. Densitometric analysis of the gel showed the effect of radiation on polypeptide constitution at radiation energy up to 7.5 kGy. Irradiation of wheat flour with 2.5 kGy have resulted in a slight increase in the molecular weight of wheat flour protein subunits. The increase of irradiation dose to 5.0 kGy has also induced an additional increase of molecular weight of protein subunits. The continuity in application of more radiation energy to a level of 7.5 kGy have resulted in the prevalence of degradation processes of all protein subunits more than the aggregation

  20. High-resolution gel electrophoresis and sodium dodecyl sulphate-agarose gel electrophoresis on urine samples for qualitative analysis of proteinuria in dogs.

    Science.gov (United States)

    Giori, Luca; Tricomi, Flavia Marcella; Zatelli, Andrea; Roura, Xavier; Paltrinieri, Saverio

    2011-07-01

    The aims of the current study were to assess whether sodium dodecyl sulphate-agarose gel electrophoresis (SDS-AGE) and high-resolution electrophoresis (HRE) can identify dogs with a urinary protein-to-creatinine ratio (UPC ratio) >0.2 and whether HRE can provide preliminary information about the type of proteinuria, using SDS-AGE as a reference method. HRE and SDS-AGE were conducted on 87 urine samples classified according to the International Renal Interest Society as non-proteinuric (NP; UPC ratio: 0.51; 40/87). SDS-AGE and HRE were positive in 14 out of 32 and 3 out of 32 NP samples and in 52 out of 55 and 40 out of 55 samples with a UPC ratio >0.20, respectively. The concordance between HRE or SDS and UPC ratio was comparable (κ = 0.59; κ = 0.55). However, specificity (90%) and positive likelihood ratio (7.76) were higher for HRE than for SDS-AGE (56% and 2.16) while sensitivity was lower (73% vs. 94%). The analysis of HRE results revealed that a percentage of albumin >41.4% and an albumin/α(1)-globulin ratio (alb/α(1) ratio) >1.46 can identify samples classified by SDS-AGE as affected by glomerular proteinuria while a percentage of α(1)-globulin >40.8% and an alb/α(1) ratio HRE could misclassify samples with a UPC ratio higher or lower than 0.20. Therefore, UPC ratio must always be determined before conducting these tests. The percentage of albumin and α(1)-globulin or the alb/α(1) ratio determined by HRE can provide preliminary information about the origin of proteinuria.

  1. An effective placental cotyledons proteins extraction method for 2D gel electrophoresis.

    Science.gov (United States)

    Tan, Niu J; Daim, Leona D J; Jamil, Amilia A M; Mohtarrudin, Norhafizah; Thilakavathy, Karuppiah

    2017-03-01

    Effective protein extraction is essential especially in producing a well-resolved proteome on 2D gels. A well-resolved placental cotyledon proteome, with good reproducibility, have allowed researchers to study the proteins underlying the physiology and pathophysiology of pregnancy. The aim of this study is to determine the best protein extraction protocol for the extraction of protein from placental cotyledons tissues for a two-dimensional gel electrophoresis (2D-GE). Based on widely used protein extraction strategies, 12 different extraction methodologies were carefully selected, which included one chemical extraction, two mechanical extraction coupled protein precipitations, and nine chemical extraction coupled protein precipitations. Extracted proteins were resolved in a one-dimensional gel electrophoresis and 2D-GE; then, it was compared with set criteria: extraction efficacy, protein resolution, reproducibility, and recovery efficiency. Our results revealed that a better profile was obtained by chemical extraction in comparison to mechanical extraction. We further compared chemical extraction coupled protein precipitation methodologies, where the DNase/lithium chloride-dense sucrose homogenization coupled dichloromethane-methanol precipitation (DNase/LiCl-DSH-D/MPE) method showed good protein extraction efficiency. This, however, was carried out with the best protein resolution and proteome reproducibility on 2D-gels. DNase/LiCl-DSH-D/MPE was efficient in the extraction of proteins from placental cotyledons tissues. In addition, this methodology could hypothetically allow the protein extraction of any tissue that contains highly abundant lipid and glycogen. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  2. [Study on the method of two dimensional polycrylamide gel electrophoresis on rat condylar chondrocyte].

    Science.gov (United States)

    Wu, Tuo-jiang; Li, Huang; Ma, Qiao-lin; Wang, Wen-mei

    2010-08-01

    To investigate the protein profile by two dimensional polycrylamide gel electrophoresis on the rat condylar chondrocyte in vitro. The third-passage chondrocytes were harvested from the mandibular condyles of 2-day-old rats in this study. The protein profile of the rat mandibular condylar chondrocytes was examined by two dimensional polycrylamide gel electrophoresis (2-DE-PAGE). The 2-DE gel maps on different pH gradients were obtained. The result of modified coomassi blue-sliver staining and sliver staining was compared using Pdquest 7.1 image analysis software. The results showed that the good protein profile of the condylar chondrocytes was obtained by standard Bio-Rad manual. The protein was mainly in the field from pH4 to pH7. The 1203±86 protein points were examined on 2-DE gel map by modified coomassi blue-sliver staining, and 1769±97 protein points was examined by sliver staining. The silver staining map showed more distinctly but higher background than modified coomassi blue-sliver staining. The protein profile of the condylar chondrocytes enriches the proteomic database and gives evidence to further proteomic research. The 2-DE map obtained by modified coomassi blue-sliver staining is more suitable for MALDI-TOF mass identification. Supported by National Natural Science Foundation of China (Grant No. C30700963), China Postdoctoral Science Foundation(Grant No.20090461088), Jiangsu Provincial Postdoctoral Science Foundation (Grant No.0802003C) and Nanjing City's Science and Technology Foundation (Grant No.200905011).

  3. Disparity between Multilocus Enzyme Electrophoresis, Microsatellite Markers and Pulsed-Field Gel Electrophoresis in epidemiological tracking of Candida albicans.

    Science.gov (United States)

    Boriollo, Marcelo Fabiano Gomes; Dias, Ricardo Antunes; Fiorini, João Evangelista; Oliveira, Nelma de Mello Silva; Spolidório, Denise Madalena Palomari; de Souza, Henrique Marques Barbosa; Figueira, Antonio Vargas de Oliveira; Pizzirani-Kleiner, Aline Aparecida

    2010-09-01

    Various molecular systems are available for epidemiological, genetic, evolutionary, taxonomic and systematic studies of innumerable fungal infections, especially those caused by the opportunistic pathogen C. albicans. A total of 75 independent oral isolates were selected in order to compare Multilocus Enzyme Electrophoresis (MLEE), Electrophoretic Karyotyping (EK) and Microsatellite Markers (Simple Sequence Repeats - SSRs), in their abilities to differentiate and group C. albicans isolates (discriminatory power), and also, to evaluate the concordance and similarity of the groups of strains determined by cluster analysis for each fingerprinting method. Isoenzyme typing was performed using eleven enzyme systems: Adh, Sdh, M1p, Mdh, Idh, Gdh, G6pdh, Asd, Cat, Po, and Lap (data previously published). The EK method consisted of chromosomal DNA separation by pulsed-field gel electrophoresis using a CHEF system. The microsatellite markers were investigated by PCR using three polymorphic loci: EF3, CDC3, and HIS3. Dendrograms were generated by the SAHN method and UPGMA algorithm based on similarity matrices (S(SM)). The discriminatory power of the three methods was over 95%, however a paired analysis among them showed a parity of 19.7-22.4% in the identification of strains. Weak correlation was also observed among the genetic similarity matrices (S(SM)(MLEE)xS(SM)(EK)xS(SM)(SSRs)). Clustering analyses showed a mean of 9+/-12.4 isolates per cluster (3.8+/-8 isolates/taxon) for MLEE, 6.2+/-4.9 isolates per cluster (4+/-4.5 isolates/taxon) for SSRs, and 4.1+/-2.3 isolates per cluster (2.6+/-2.3 isolates/taxon) for EK. A total of 45 (13%), 39 (11.2%), 5 (1.4%) and 3 (0.9%) clusters pairs from 347 showed similarity (S(J)) of 0.1-10%, 10.1-20%, 20.1-30% and 30.1-40%, respectively. Clinical and molecular epidemiological correlation involving the opportunistic pathogen C. albicans may be attributed dependently of each method of genotyping (i.e., MLEE, EK, and SSRs) supplemented

  4. A Robust Identification of the Protein Standard Bands in Two-Dimensional Electrophoresis Gel Images

    Directory of Open Access Journals (Sweden)

    Serackis Artūras

    2017-12-01

    Full Text Available The aim of the investigation presented in this paper was to develop a software-based assistant for the protein analysis workflow. The prior characterization of the unknown protein in two-dimensional electrophoresis gel images is performed according to the molecular weight and isoelectric point of each protein spot estimated from the gel image before further sequence analysis by mass spectrometry. The paper presents a method for automatic and robust identification of the protein standard band in a two-dimensional gel image. In addition, the method introduces the identification of the positions of the markers, prepared by using pre-selected proteins with known molecular mass. The robustness of the method was achieved by using special validation rules in the proposed original algorithms. In addition, a self-organizing map-based decision support algorithm is proposed, which takes Gabor coefficients as image features and searches for the differences in preselected vertical image bars. The experimental investigation proved the good performance of the new algorithms included into the proposed method. The detection of the protein standard markers works without modification of algorithm parameters on two-dimensional gel images obtained by using different staining and destaining procedures, which results in different average levels of intensity in the images.

  5. Radiobiological study on DNA strand breaks and repair using single cell gel electrophoresis

    International Nuclear Information System (INIS)

    Ikushima, Takaji

    1994-01-01

    Single cell gel electrophoresis (SCGE) provides a novel method to measure DNA damage in individual cells and more importantly, to assess heterogeneity in response within a mixed population of cells. Cells embedded in agarose are lysed, subjected to electrophoresis, stained with a fluorescent DNA-specific dye, and viewed under a fluorescence microscope. Damaged cells display 'comets', broken DNA migrating farther to the anode in the electric field. We have previously used this technique to quantify DNA damage induced by moderate doses of low and high LET radiations in cultured Chinese hamster cells. The assay has been optimized in terms of lysing and electrophoresis conditions, and applied to analyse the DNA strand breaks, their repair kinetics and heterogeneity in response in individual Chinese hamster cells exposed to gamma-rays, and to KUR thermal neutrons with and without 10 B or to KEK PF monochromatic soft X-rays as well as to a radio-mimetic agent, neocarzinostatin. The DNA double-strand breaks induced by boron-neutron captured reactions were repaired at a slower rate, but a heterogeneity in response might not contribute to the difference. The neocarzinostatin-induced DNA damage were efficiently repaired in a dose-dependent fashion. The initial amount of gamma-ray induced DNA double-strand breaks was not significantly altered in cells pre-exposed to very low adapting dose. (author)

  6. Comparative analyses of amplicon migration behavior in differing denaturing gradient gel electrophoresis (DGGE) systems

    Science.gov (United States)

    Thornhill, D. J.; Kemp, D. W.; Sampayo, E. M.; Schmidt, G. W.

    2010-03-01

    Denaturing gradient gel electrophoresis (DGGE) is commonly utilized to identify and quantify microbial diversity, but the conditions required for different electrophoretic systems to yield equivalent results and optimal resolution have not been assessed. Herein, the influence of different DGGE system configuration parameters on microbial diversity estimates was tested using Symbiodinium, a group of marine eukaryotic microbes that are important constituents of coral reef ecosystems. To accomplish this, bacterial clone libraries were constructed and sequenced from cultured isolates of Symbiodinium for the ribosomal DNA internal transcribed spacer 2 (ITS2) region. From these, 15 clones were subjected to PCR with a GC clamped primer set for DGGE analyses. Migration behaviors of the resulting amplicons were analyzed using a range of conditions, including variation in the composition of the denaturing gradient, electrophoresis time, and applied voltage. All tests were conducted in parallel on two commercial DGGE systems, a C.B.S. Scientific DGGE-2001, and the Bio-Rad DCode system. In this context, identical nucleotide fragments exhibited differing migration behaviors depending on the model of apparatus utilized, with fragments denaturing at a lower gradient concentration and applied voltage on the Bio-Rad DCode system than on the C.B.S. Scientific DGGE-2001 system. Although equivalent PCR-DGGE profiles could be achieved with both brands of DGGE system, the composition of the denaturing gradient and application of electrophoresis time × voltage must be appropriately optimized to achieve congruent results across platforms.

  7. Application of the Single Cell Gel Electrophoresis (SCGE) Assay to Genotoxicity Evaluation in Plants and Animals

    International Nuclear Information System (INIS)

    Kim, Jin Kyu

    2007-10-01

    Application of the Single Cell Gel Electrophoresis (SCGE) Assay to Genotoxicity Evaluation in Plants and Animals. Recently, the importance of ionizing radiation and chemicals has been recognized since radio- and chemical therapy is directly related to the control of various diseases such as cancer. Radiation and the chemicals can cause biological damages while they have great applicability. It is of necessity to analyze rapidly, easily and accurately the biological effects, especially DNA damage due to those factors. Recently SCGE (single cell gel electrophoresis assay, alias comet assay) has been developed for the efficient evaluation of DNA damage. In this report, the comprehensive review will be given on the rationale, the technical applications and the advantages and shortcomings of SCGE assay. This method can be directly applied to study on toxicity, cancer, and aging in terms of the evaluation of DNA damages due to radiation and chemicals on human cellular level. It is also suggested that comet assay be used for testing genotoxicity of suspected substances, detecting irradiated foods, screening radioprotective candidates, and studying DNA repair process in various biological systems

  8. Two-dimensional gel electrophoresis data for proteomic profiling of Sporothrix yeast cells

    Directory of Open Access Journals (Sweden)

    Anderson Messias Rodrigues

    2015-03-01

    Full Text Available Sporotrichosis is a chronic infection of the skin and subcutaneous tissues of human and other mammals caused by a complex of cryptic dimorphic fungi in the plant-associated order Ophiostomatales. With major differences between routes of transmission, Sporothrix infections are emerging as new threat in tropical and subtropical areas, particularly in form of outbreaks. The mechanisms underlying the pathogenesis and invasion of Sporothrix spp. are still poorly understood and many virulence factors remain unidentified. In this scenario, a global analysis of proteins expressed by clinical Sporothrix species combined with the identification of seroreactive proteins is overdue. Optimization of sample preparation and electrophoresis conditions are key steps toward reproducibility of gel-based proteomics assays. We provide the data generated using an efficient protocol of protein extraction for rapid and large-scale proteome analysis using two-dimensional gel electrophoresis. The protocol was established and optimized for pathogenic and non-pathogenic Sporothrix spp. including Sporothrix brasiliensis (CBS 132990, Sporothrix schenckii sensu stricto (CBS 132974, Sporothrix globosa (CBS 132922, and Sporothrix mexicana (CBS 120341. The data, supplied in this article, are related to the research article entitled “Immunoproteomic analysis reveals a convergent humoral response signature in the Sporothrix schenckii complex” (Rodrigues et al., 2014 [1].

  9. Application of the Single Cell Gel Electrophoresis (SCGE) Assay to Genotoxicity Evaluation in Plants and Animals

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Jin Kyu

    2007-10-15

    Application of the Single Cell Gel Electrophoresis (SCGE) Assay to Genotoxicity Evaluation in Plants and Animals. Recently, the importance of ionizing radiation and chemicals has been recognized since radio- and chemical therapy is directly related to the control of various diseases such as cancer. Radiation and the chemicals can cause biological damages while they have great applicability. It is of necessity to analyze rapidly, easily and accurately the biological effects, especially DNA damage due to those factors. Recently SCGE (single cell gel electrophoresis assay, alias comet assay) has been developed for the efficient evaluation of DNA damage. In this report, the comprehensive review will be given on the rationale, the technical applications and the advantages and shortcomings of SCGE assay. This method can be directly applied to study on toxicity, cancer, and aging in terms of the evaluation of DNA damages due to radiation and chemicals on human cellular level. It is also suggested that comet assay be used for testing genotoxicity of suspected substances, detecting irradiated foods, screening radioprotective candidates, and studying DNA repair process in various biological systems.

  10. Effect of gamma-irradiation on cereal DNA investigated by pulsed-field gel electrophoresis

    International Nuclear Information System (INIS)

    Kawamura, Yoko; Miura, Aya; Imura, Hiromi; Yamada, Takashi; Saito, Yukio

    1996-01-01

    The effects of gamma-irradiation on the DNA of corn, soybean and wheat were investigated using a pulsed-field gel electrophoresis technique. In order to avoid strand breaks during the DNA extracting steps, protoplasts prepared from seeds were embedded in agarose plugs and the DNA was purified by the digesting membranes and proteins. Pulsed-field gel electrophoresis can separate large DNA strands of about a few Mb in length. The DNA from unirradiated corn, soybean and wheat had mainly 3 fragments, about 6Mb(Fr.1), 5Mb(Fr.2), a few hundred kb(Fr.3) and so on. After gamma-irradiation, Fr.1 and Fr.2 had decreased depend on irradiation dose. The Fr.4(about 200 kb) of corn and Fr.3 of soybean DNA increased while Fr.3 of wheat did not increase under 10 kGy irradiation, however, the Fr.3 of all samples and the Fr.4 of corn decreased by over 10 kGy irradiation. It can be assumed that the large DNA strands were broken into smaller strands which increased at low irradiation doses, whereas both large and small DNA strands were broken down at higher irradiation doses. The Fr.6(2.5Mb) and Fr.7(1.5Mb) appeared in irradiated wheat DNA. (author)

  11. Evaluation of agarose gel electrophoresis for characterization of silver nanoparticles in industrial products.

    Science.gov (United States)

    Jimenez, Maria S; Luque-Alled, Jose M; Gomez, Teresa; Castillo, Juan R

    2016-05-01

    Agarose gel electrophoresis (AGE) has been used extensively for characterization of pure nanomaterials or mixtures of pure nanomaterials. We have evaluated the use of AGE for characterization of Ag nanoparticles (NPs) in an industrial product (described as strong antiseptic). Influence of different stabilizing agents (PEG, SDS, and sodium dodecylbenzenesulfonate), buffers (TBE and Tris Glycine), and functionalizing agents (mercaptosuccinic acid (TMA) and proteins) has been investigated for the characterization of AgNPs in the industrial product using different sizes-AgNPs standards. The use of 1% SDS, 0.1% TMA, and Tris Glycine in gel, electrophoresis buffer and loading buffer led to the different sizes-AgNPs standards moved according to their size/charge ratio (obtaining a linear relationship between apparent mobility and mean diameter). After using SDS and TMA, the behavior of the AgNPs in the industrial product (containing a casein matrix) was completely different, being not possible their size characterization. However we demonstrated that AGE with LA-ICP-MS detection is an alternative method to confirm the protein corona formation between the industrial product and two proteins (BSA and transferrin) maintaining NPs-protein binding (what is not possible using SDS-PAGE). © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Impact of quenching failure of Cy dyes in differential gel electrophoresis.

    Directory of Open Access Journals (Sweden)

    Weiqun Wang

    Full Text Available BACKGROUND: Differential gel electrophoresis (DIGE is a technology widely used for protein expression analysis. It is based on labelling with fluorescent Cy dyes. In comparative fluorescence gel electrophoresis experiments, however, unspecific labelling using N-hydroxy-succinimide-ester-based labelling protocols was recently detected. Cross-talk was observed due to failure of the quenching process. Here, the impact of this effect for DIGE experiments was investigated. METHODOLOGY/PRINCIPAL FINDINGS: Experiments to test quenching efficiency were performed in replicate using Escherichia coli lysate. Parameters such as the amount of dye and quencher were varied. Labelling and quenching were reversed in one experiment. Differences in protein spot volumes due to limited quenching were determined. For some spots twice the volume was detected underscoring the importance of proper control of silencing of active dye. CONCLUSIONS/SIGNIFICANCE: It could be demonstrated that uncontrolled labelling increased protein spot volume, even doubling it in some cases. Moreover, proteins responded differently to the protocol. Such unpredictable and unspecific processes are not acceptable in protein regulation studies so that it is necessary to validate the correct amount of quencher for individual samples before the DIGE experiment is performed. Increase of the concentration of lysine, which is used as quencher, from 10 mM to 2500 mM, was sufficient to silence the dye. Alternatively, active dye molecules can be removed by filtration.

  13. Agarose-gel electrophoresis for the quality assurance and purity of heparin formulations.

    Science.gov (United States)

    Volpi, Nicola; Buzzega, Dania

    2012-01-01

    The adulteration of raw heparin (Hep) with a synthetic oversulfated chondroitin sulfate (OSCS) not found in nature produced in 2007-2008 a global crisis giving rise to the development of additional, new and specific methods for its quality assurance and purity. In this study, a simple and sensitive agarose-gel electrophoresis method has been developed for the visualization of OSCS in Hep samples along with other natural glycosaminoglycans possibly present as "process-related impurities", in particular dermatan sulfate (DS) and chondroitin sulfate (CS). Agarose-gel electrophoresis under non-conventional conditions is able to separate OSCS from Hep with its two components, the slow-moving and fast-moving species, DS and CS by performing separation for 15 h (overnight) and under high voltage (100 mA, ∼200 V). Densitometric scanning enabled us to calculate a limit of detection of ∼0.5 μg OSCS with a linear behaviour from 0.1 to 5 μg, comparable to CS/DS. Contaminated samples from Hep manufacturers were analyzed and quantitative data were found comparable to previous studies. Due to its capacity to process many samples in a single run and to the equipment commonly available in laboratories, this analytical method would be suitable for the identification and quantification of contamination by other polysaccharides, in particular OSCS and DS, within Hep preparations and formulations. Copyright © 2012 Elsevier B.V. All rights reserved.

  14. Non-aqueous capillary electrophoresis with red light emitting diode absorbance detection for the analysis of basic dyes.

    Science.gov (United States)

    Fakhari, Ali Reza; Breadmore, Michael C; Macka, Miroslav; Haddad, Paul R

    2006-11-24

    Non-aqueous capillary electrophoresis was evaluated for the separation of five hydrophobic basic blue dyes for application in forensic dye analysis. The use of a red light emitting diode as a high intensity, low-noise light source provided sensitive detection of the blue dyes while also allowing the evaluation of solvents that absorb strongly in the UV region. Excellent peak shapes and separation selectivity were obtained in methanol, ethanol, acetonitrile and dimethylsulfoxide, however water, tetrahydrofuran, dimethylformamide and acetone were unsuitable as solvents due to poor peak shapes and a lack of sensitivity, most likely due to adsorption onto the capillary wall. Due to the known compatibility of methanol with capillary electrophoresis-mass spectrometry, this solvent was examined further with the relative acidity/basicity of the electrolyte being optimised with an artificial neural network. The optimised method was examined for the separation of ink samples from 6 fibre tip and 2 ball point blue or black pens and showed that a unique migration time for the main dye component in seven of the eight pens could be obtained.

  15. Optimization of a pulsed-field gel electrophoresis for molecular typing of Proteus mirabilis

    Directory of Open Access Journals (Sweden)

    Alper Karagöz

    2013-09-01

    Full Text Available Objective: For the detection of outbreaks caused byProteus mirabilis, strains clonal relations are determinedmethods as “pulsed-field gel electrophoresis (PFGE”.The aim of this study was optimization of a pulsed-fieldgel electrophoresis for molecular typing of P. mirabilis.Methods: In this study, PFGE’ protocol is optimized foruse in molecular typing of P. mirabilis. Phylogenetic analyzesof strains were evaluated with Bionumerics softwaresystem (version 6.01; Applied Maths, Sint-Martens-Latem, Belgium.Results: This protocol compared with Gram-negativebacteria PFGE protocols, NotI enzyme is suitable for thisbacterium. Electrophoresis conditions should be revealedas; - block 1: initial pulse duration 1 sec, ending pulseduration 30 sec, striking angle 120°, the current 6 V/cm2,temperature 14°C, time 8 hours; - block 2: initial pulseduration 30 sec, ending pulse duration 70 sec, strikingangle 120°, the current 6 V/cm2, temperature 14°C, time16 hours; - TBE, pH=8.4.Conclusion: P. mirabilis strains were typed by PFGE andBionumerics analysis program were determined clonal relationships.The procedure was simple, reproducible andsuitable for these bacteria. Also it was evaluated, becauseof reducing time, the solution volumes and enzymes canbe economically. Outbreaks of nosocomial infections dueto bacteria studied assessment and the potential to provideuseful information about the degree of prevalence.This optimized protocol is allowed different centers’ PFGEresults to compare with other laboratories results. J ClinExp Invest 2013; 4 (3: 306-312Key words: Proteus mirabilis, molecular typing, pulsedfieldgel electrophoresis.

  16. Determination of effective charges and ionic mobilities of polycationic antimicrobial peptides by capillary isotachophoresis and capillary zone electrophoresis

    Czech Academy of Sciences Publication Activity Database

    Tůmová, Tereza; Monincová, Lenka; Nešuta, Ondřej; Čeřovský, Václav; Kašička, Václav

    2017-01-01

    Roč. 38, č. 16 (2017), s. 2018-2024 ISSN 0173-0835 R&D Projects: GA ČR(CZ) GA15-01948S Institutional support: RVO:61388963 Keywords : antimicrobial peptides * capillary isotachophoresis * counterion condensation * effective charge * ionic mobility Subject RIV: CB - Analytical Chemistry, Separation OBOR OECD: Analytical chemistry Impact factor: 2.744, year: 2016

  17. 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

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

    Directory of Open Access Journals (Sweden)

    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.

  19. Design and evaluation of capillary coupled with optical fiber light-emitting diode induced fluorescence detection for capillary electrophoresis.

    Science.gov (United States)

    Ji, Hongyun; Li, Meng; Guo, Lihong; Yuan, Hongyan; Wang, Chunling; Xiao, Dan

    2013-09-01

    A new detector, capillary coupled with optical fiber LED-induced fluorescence detector (CCOF-LED-IFD, using CCOF for short), is introduced for CE. The strategy of the present work was that the optical fiber and separation capillary were, in the parallel direction, fastened in a fixation capillary with larger inner diameter. By employing larger inner diameter, the fixation capillary allowed the large diameter of the optical fiber to be inserted into it. By transmitting an enhanced excitation light through the optical fiber, the detection sensitivity was improved. The advantages of the CCOF-CE system were validated by the detection of riboflavin, and the results were compared to those obtained by the in-capillary common optical fiber LED-induced fluorescence detector (IC-COF-LED-IFD, using COF for short). The LODs of CCOF-CE and COF-CE were 0.29 nM and 11.0 nM (S/N = 3), respectively. The intraday (n = 6) repeatability and interday (n = 6) reproducibility of migration time and corresponding peak area for both types of CE were all less than 1.10 and 3.30%, respectively. The accuracy of the proposed method was judged by employing standard addition method, and recoveries obtained were in the range of 98.0-102.4%. The results indicated that the sensitivity of the proposed system was largely improved, and that its reproducibility and accuracy were satisfactory. The proposed system was successfully applied to separate and determine riboflavin in real sample. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Electrophoretic analysis of proteins from Mycoplasma hominis strains detected by SDS-PAGE, two-dimensional gel electrophoresis and immunoblotting

    DEFF Research Database (Denmark)

    Andersen, H; Birkelund, Svend; Christiansen, Gunna

    1987-01-01

    The proteins of 14 strains of Mycoplasma hominis were compared by SDS-PAGE in gradient gels, by two-dimensional (2D) gel electrophoresis of extracts of 35S-labelled cells and by immunoblot analysis of cell proteins. The strains examined included the M. hominis type strain PG21 and 13 others...... isolated variously from genital tract, mouth, blood, upper urinary tract and a wound. These 14 strains shared 76-99% of proteins in SDS-gradient gel analysis and 41-72% in the 2D gels. As expected, the immunoblot analysis likewise revealed the existence of an extensive common protein pattern in M. hominis...