bluuebunny/crossref_metadata_2025_split huggingface dataset:
Capillary electrophoresis of peptides using rectangular and cylindrical geometries: A comparative study
Authors: Cifuentes Alejandro , Poppe Hans 
Published: May 2005

AbstractA study on the use of rectangular columns in capillary zone electrophoresis (CZE) is presented. The equations controlling the hydrodynamic flow and the plate height in rectangular capillaries are deduced. Several experiments were carried out in order to test the validity of these equations. Also, the possibilities of these columns in terms of thermal dissipation, efficiency, analysis speed and sample capacity were studied and compared with those for cylindrical tubing. The results from both rectangular and cylindrical columns are compared to each other, employing the parameters mentioned above, and also with those predicted by theory. The advantages, i.e. better heat dissipation, higher speed and larger sample capacity, and drawbacks, i.e. high fragility and incompatibility with some capillary electrophoresis instruments, of employing rectangular geometries in CZE are discussed.
An expanded system of restriction landmark genomic scanning (RLGS Ver. 1.8)
Authors: Okazaki Yasushi , Okuizumi Hisato , Sasaki Nobuya , Ohsumi Tomoya , Kuromitsu Junro , Hirota Norifumi , Muramatsu Masami , Hayashizaki Yoshihide 
Published: May 2005

AbstractThe restriction landmark genomic scanning (RLGS) method is a high‐speed genome scanning system which is based on the concept that restriction enzyme sites can be used as landmarks throughout the genome. It employs direct end‐labeling of the genomic DNA digested with a rare‐cutting restriction enzyme, followed by high‐resolutional two‐dimensional electrophoresis. Recently, this system was further developed to lower cost and to simplify the procedure. This paper reviews the RLGS principle and the breakthroughs enabling its further development. Also presented is the precise protocol of the newest version (RLGS Ver. 1.8) that offers cost effectiveness and an expanded production system. Finally, the advantages of this new RLGS method and prospects for its widespread application are discussed.
Theory of capillary electrophoretic separations of DNA‐polymer complexes
Authors: Hubert Sylvain J. , Slater Gary W. 
Published: May 2005

AbstractElectrophoretic separation of DNA molecules normally requires the use of an anticonvection, sieving polymer matrix such as a gel or an entangled polymer solution. Recently, it has been suggested that free‐solution separation could be achieved in a capillary if an electrically neutral, friction‐generating molecule is attached to the DNA molecules before electrophoresis is carried out. The electrophoretic mobilities are then predicted to be very large and the resulting separation is expected to yield excellent resolution. The size‐dependence of the electrophoretic mobility is attributed to longer DNA molecules pulling the neutral molecule with a larger electric force, thus eluting earlier than shorter DNA molecules. In this article, we focus on the particular case where one attaches an uncharged, flexible polymer to the end of the DNA. Our self‐consistent model takes into account the deformation and the hydrodynamic resistance of the polymer in the flow. We find various regimes, depending on the intensity of the electric field and the length of the polymer. The most favorable conditions for high‐resolution separation of DNA are described.
Application of chemometric experimental designs in capillary electrophoresis: A review
Authors: Altria Kevin D. , Clark Brian J. , Filbey Sharon D. , Kelly Martin A. , Rudd David R. 
Published: May 2005

AbstractVarious chemometric experimental designs have been employed for the optimisation of capillary electrophoresis (CE) methods. Similar designs have been utilised in the assessment of the robustness of CE methods. The designs employed include central composites, fractional factorials, Plackett‐Burman, simplex and overlapping‐resolution mapping. Optimisation studies have largely concentrated on the use of these designs on selection of the optimal electrolyte composition. The robustness testing studies performed have involved the use of screening designs to identify the critical parameters affecting responses such as migration times and resolution. Further designs such as central composites have then been employed to set method limits following robustness studies. It is concluded that the use of experimental designs and statistical data evaluation in conjunction with personal computer‐controlled CE autosamplers and instruments are of great benefit in the optimisation and robustness evaluation of CE methods.
Expression of cDNA clones by coupled in vitro transcription/translation and transfection into COS‐1 cells: Protein mapping in two‐dimensional gels
Authors: Madsen Peder , Gromov Pavel , Celis Julio E. 
Published: May 2005

AbstractWe present two procedures that can be used to map proteins in two‐dimensional gels if the cDNA is at hand. The first procedure, which is illustrated with the expression of cDNAs encoding the fatty acid binding protein 5 (FABP 5), psoriasin and stratifin, makes use of the in vitro transcription/translation assay marketed by Promega. The procedure is simple and allows the mapping of the primary translation product in a very short time. The second method – which faithfully reproduces post‐translational modifications – is based on the expression of cDNAs transfected into COS‐1 cells using a eukaryotic expression plasmid. This procedure is illustrated with the expression of cDNAs encoding Ha‐ras p21 and rab 11, two small GTP‐binding proteins known to undergo complex modifications.
Uses of digital image analysis in electrophoresis
Authors: Horgan Graham W. , Glasbey Chris A. 
Published: May 2005

AbstractThe information in electrophoretic gels can be extracted visually, or by using specialised computer hardware and software which treat the gels as digital images. We discuss issues which arise in applying digital image analysis to electrophoresis, and comment on the range of computer packages available. We also illustrate how image analysis can make gel interpretation easier and more reliable. Methods covered include gel registration, and warping, mathematical morphology and deconvolution to sharpen images.
The use of sodium dodecyl sulphate‐polyacrylamide gel electrophoresis to confirm the presence of salmon (Salmo salar) eggs in an illegal fish bait
Authors: Scobbie Alan E. , Mackie Ian M. 
Published: May 2005

AbstractSalmon roe was identified as the major component of a suspected illegal fish attractant using sodium dodecyl sulphate‐polyacrylamide gel electrophoresis to analyse protein extracts prepared from various salmonid eggs and a suspected illegal bait.
Identification of myocardial proteins from two‐dimensional gels by peptide mass fingerprinting
Authors: Sutton Chris W. , Pemberton Kay S. , Cottrell John S. , Corbett Joseph M. , Wheeler Colin H. , Dunn Michael J. , Pappin Darryl J. 
Published: May 2005

AbstractTwo‐dimensional gels offer a powerful method for separating complex protein mixtures, but subsequent methods for analysing individual components, such as protein sequencing and Western immunoblotting, are laborious and slow. The identification of proteins can be accelerated by using a combination of protease digestion and matrix assisted laser desorption‐mass spectrometry (MALDI‐MS). The peptide mass spectrum of a protein represents a unique fingerprint determined by the amino acid sequence and the cleavage properties of the protease. Software has been developed so that peptide masses can be used to search a mass‐based peptide database generated from established protein sequence databases. A list of the closest matching proteins is produced to allow identification of the sample. The strategy was applied to 52 protein spots from human myocardial tissue separated by two‐dimensional electrophoresis (2‐DE) gels and analysed blind. Conditions for optimal trypsin digestion of proteins electroblotted onto polyvinylidene difluoride (PVDF) membranes are described. Mass data were generated from both Coomassie Brilliant Blue and sulforhodamine B‐stained proteins, though the former required destaining prior to digestion. Alkylation of cysteine and oxidation of methionine were significant modifications that influenced the successful identification of a protein spot. Examples are presented to illustrate the advantages and disadvantages of this approach.
Detection of picornavirus genomic and template RNA strands by a novel semi‐nested polymerase chain reaction‐technique and agarose gel electrophoresis
Authors: Gow John , Cash Philip , Behan Wilhelmina , McGarry Frances , Simpson Kathleen , Behan Peter 
Published: May 2005

AbstractAttempts to evaluate the relative levels of enteroviral genomic and template RNA strands in small biopsy tissue samples from patients have yielded ambiguous data, largely due to the limited amount of RNA available. A novel seminested polymerase chain reaction (PCR) technique was developed to enable RNA levels to be examined more accurately. PCR products were visualised by horizontal agarose gel electrophoresis. This technique was demonstrated on linear single‐stranded plasmid DNA; viral RNA isolated from a human rhabdomyosarcoma (RD) cell line persistently infected with a mutated coxsackie B5 virus (piRD) and two cell lines, RD and HEp2 cells, acutely infected with a wild‐type clinical isolate of coxsackie B5 virus.
Sequence dependent migration behavior of double‐stranded DNA in capillary electrophoresis
Authors: Berka Jan , Pariat Yvan F. , Müller Odilo , Hebenbrock Kirstin , Heiger David N. , Foret František , Karger Barry L. 
Published: May 2005

AbstractCapillary electrophoresis (CE) with a replaceable linear polyacrylamide (LPA) sieving matrix was used to examine sequence‐dependent migration of double‐stranded DNA fragments. It has been found that DNA conformational effects were significant under high electric field separations, especially using high resolution matrices. Compared to linear DNA‐ladder standards, both anomalously slow and rapid DNA fragments were observed, with the degree of anomalous migration depending on the electric field strength, polymer concentration, column temperature, and background electrolyte (denaturants, sodium and magnesium ions, DNA‐intercalating dyes). By selecting a combination of electrophoretic conditions (e.g. 3% T LPA, elevated capillary temperature, lower electric field strength and addition of DNA intercalating dyes), molecular weight dependent separations were closely restoredPresented in part at HPCE '94, San Diego, CA, February, 1994.
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Analysis of heat and cold shock proteins in Listeria by two‐dimensional electrophoresis
Authors: Phan‐Thanh Luu , Gormon Thierry 
Published: May 2005

AbstractThe proteins induced by heat and cold shock in Listeria monocytogenes (pathogenic for humans) and L. innocua (nonpathogenic) strains were analyzed by two‐dimensional (2‐D) electrophoresis with the help of a computerized 2‐D gel analysis system. Heat (49°C) and cold (4°C) shock repressed roughly half the number of proteins synthesized at normal temperature (25°C) and decreased the level of numerous other proteins. Conversely, the synthesis of a great number of proteins was enhanced and novel proteins appeared upon temperature stress. There were more proteins induced in the L. monocytogenes strain than in the L. innocua strain. Each stress induced a set of specific proteins. There was overlap between these sets of proteins induced by heat and cold shock. Furthermore, a number of heat or cold shock proteins were found to be induced in both Listeria species and by both heat and cold shock in both species. The induction by heat shock was more intense than that by cold shock. The most strongly induced common stress protein of Listeria had a molecular mass of 17.6 kDa and an isoelectric point of 5.1.
Direct determination of amino acids by capillary electrophoresis/electrochemistry using a copper microelectrode and zwitterionic buffers
Authors: Zhou Jianxun , Lunte Susan M. 
Published: May 2005

AbstractSixteeen free amino acids were separated by capillary electrophoresis (CE) using zwitterionic buffers of pH 9.8 and detected amperometrically using a copper microelectrode. Low conductivity buffers make possible the use of higher buffer concentrations. This depresses the electroosmotic flow, leading to a better separation of the amino acids compared to that obtained using strong electrolytes. The use of zwitterionic buffers also minimizes the amount of Joule heating in the capillary, leading to more efficient separations. Perhaps most importantly for electrochemical (EC) detection, these buffers reduce the noise due to high separation current at the EC detector. The increased signal‐to‐noise ratio (S/N) results in much better detection limits for amino acids than those reported previously for capillary electrophoresis/electrochemistry (CEEC) with a copper electrode. Detection limits ranged from 10 to 400 nm, and the response was linear over three orders of magnitude for most of the amino acids. The system demonstrated good long‐term stability and reproducibility with a relative standard deviation (RSD) of less than 5% for both the migration time and peak current (n = 20). This method was employed for the determination of amino acids in urine and in brain microdialysate samples.
High performance capillary electrophoresis of casein phosphopeptides containing 2‐5 phosphoseryl residues: Relationship between absolute electrophoretic mobility and peptide charge and size
Authors: Adamson Nicholas J. , Reynolds Eric C. 
Published: May 2005

AbstractMultiple phosphoseryl‐containing peptides from casein containing the cluster sequence ‐Ser(P)‐Ser(P)‐Ser(P)‐Glu‐Glu‐ stabilize amorphous calcium phosphate at neutral and alkaline pH and have been shown to be anticariogenic in various in vitro, animal and human experiments. In an approach to obtain insight into the structure and function of these peptides, we previously developed a method for their analysis using high‐performance capillary electrophoresis (HPCE). A linear relationship was obtained between absolute electrophoretic mobility and q/M2/3 where q is the net negative charge of the peptide calculated using relevant pKa values and M is the molecular mass. M2/3 is a measure of the surface area of a sphere that has a volume proportional to the M of the peptide and relates to the frictional drag exerted on the peptide during electrophoretic migration. To examine this relationship further, we have analyzed a range of high‐performance liquid chromatography (HPLC)‐purified cluster, di‐ and triphosphorylated peptides from casein enzymic (trypsin, alcalase and pancreatin) digests using HPCE and attempted to relate their absolute electrophoretic mobility to various models incorporating peptide size and charge. Absolute electrophoretic mobility of casein phosphopeptides containing the cluster sequence‐Ser(P)‐Ser(P)‐Ser(P)‐Glu‐Glu‐ was most closely correlated with q/M2/3. Di‐ and triphosphorylated peptide absolute electrophoretic mobility correlated with both q/M2/3 and In(q+1)/n0.43. However, for both the q/M2/3 and In(q+1)/n0.43 relationships with absolute electrophoretic mobility, the di‐ and triphosphorylated peptides formed a separate linear relationship to that of the cluster peptides. From these relationships, a di‐ or triphosphorylated peptide exhibited a greater absolute electrophoretic mobility than a corresponding cluster peptide with the same q/M2/3 or In(q+1)/n0.43 value. This implies that the cluster peptides have a reduced effective net negative charge due to an electrical double layer effect and/or a larger hydrodynamic volume or a more extended structure relative to the di‐ and triphosphorylated peptides which is associated with a greater frictional drag during electrophoretic migration.
Separation of eleven angiotensin II analogs by capillary electrophoresis with a nonionic surfactant in acidic media
Authors: Matsubara Norio , Koezuka Kiyohiro , Terabe Shigeru 
Published: May 2005

AbstractEleven angiotension II analogs of same chain length were separated by capillary electrophoresis at pH 2.0 with 200 mM Tween 20. All compounds except one pair of angiotensin II ([Sar1, Gly8]‐ and [Sar1, Val5, Ala8]‐angiotensin II) were baseline‐separated, even in the case of peptides with about the same total charge. The migration order of the angiotensin II analogs were determined by the hydrophobicity of the amino acid as long as the difference between amino acids of two peptides is the conservative change. From the study of pH dependency of the separation of these peptides, it was found that the conditions of a low electroosmotic flow under a low pH is effective for the separation of similar peptides.
A simplified silver diammine method for the staining of nucleic acids in polyacrylamide gels
Authors: Vari Frank , Bell Kevin 
Published: May 2005

AbstractHigh sensitivity and low background, which are the significant features of the procedure of Johansson and Skoog (J. Biochem. Biophys. Methods. 1987, 14, (Suppl.) 33) for silver staining of nucleic acids in polyacrylamide gels, have been improved by excluding ethanol from all solutions and increasing the length of washing steps after sensitisation and staining. Including a fixation step before sensitisation increased the contrast of the bands even at high concentration of nucleic acid, producing a shift in the tone of colour from black to grey instead of gold to yellow bands. This technique is superior in terms of sensitivity and background to a number of other silver staining protocols, three which utilise silver diammine and three which utilise silver nitrate ions.
Reproducibility of mobility in gel electrophoresis
Authors: Zakharov Sergey F. , Chang Huang‐Tung , Chrambach Andreas 
Published: May 2005

AbstractThe quantitative exploitation of gel electrophoresis to yield molecular and gel fiber properties rests on the assumption that mobility is characteristic of the macromolecule migrating as a band and is a physical constant for any system defined by pH, ionic strength and temperature. This assumption has not been tested intra‐experimentally in previous literature. With the commercial introduction of automated gel electrophoresis apparatus, the collection of multiple mobility data during a single run without additional expense of labor has made it possible to test the assumption. As a start, we undertook that test for three proteins and their sodium dodecyl sulfate (SDS) derivatives, in agarose and polyacrylamide gel electrophoresis, various field strengths, continuous and discontinuous buffers, as well as intra‐ and interexperimentally. It was found that in agarose gel electrophoresis conducted in a single buffer, the standard deviation of mobility over a wide concentration range ranges intra‐experimentally from 0.2 to 1.3% for two globular proteins and 1.4 to 5.3% for the same proteins derivatized with SDS. Interexperimentally, it was 3% in the single case tested to date. The standard deviation in polyacrylamide appears to be higher, varies in inverse relation to the mobility value, i.e. increases with gel concentration in the range of 11 to 19%T, and varies substantially between the two SDS‐proteins investigated. Mobility in a discontinuous buffer system decreases continuously due to the decreasing leading phase/trailing phase ratio along the migration path. The decrease is sharpest in the “nonrestrictive” stacking gel.
Investigation of wool protein heterogeneity using two‐dimensional electrophoresis with immobilised pH gradients
Authors: Herbert Ben R. , Chapman Anna L. P. , Rankin Douglas A. 
Published: May 2005

AbstractThe heterogeneity of intermediate filament proteins (IFP) from wool has been investigated using two‐dimensional polyacrylamide gel electrophoresis with immobilised pH gradients in the first dimension. The charge heterogeneity has been confirmed with some of the IFP separated as a string of spots with similar molecular weight, but markedly different in isoelectric point (pI). The molecular weight and pI distribution of the string of IFP changed after treatment with alkaline phosphatase, indicating that some of the heterogeneity is caused by phosphorylation.
Adherence of oral microorganisms to human parotid salivary proteins
Authors: Newman Frances , Beeley Josie A. , MacFarlane T. Wallace 
Published: May 2005

AbstractBacterial colonisation of oral surfaces by microorganisms may be dependent on their interaction with specific host receptor molecules. Primary oral colonisers are known to remove specific proteins from parotid saliva. The aim of this study was to determine whether these interactions facilitate microbial attachment to a surface and hence identify specific salivary components as putative host receptor molecules. Parotid saliva was resolved by sodium dodecyl sulphate‐polyacrylamide gel electrophoresis (SDS‐PAGE) and then electroblotted onto nitrocellulose membranes. Suspensions of fluorescently labelled microoganisms were incubated with the blots and salivary components with adherent bacteria identified as fluorescent bands under ultraviolet (UV) transillumination. Species of streptococci known to be early colonisers of the clean tooth surface were found to adhere specifically to certain salivary proteins, especially to basic proline‐rich proteins (PRPs). Polymorphic variations in these patterns could form the basis of differences in oral microflora, susceptibility to oral infections and consequent disease.
Separation of 1‐aminopyrene‐3,6,8‐trisulfonate‐labeled asparagine‐linked fetuin glycans by capillary gel electrophoresis
Authors: Guttman András , Chen Fu‐Tai A. , Evangelista Ramon A. 
Published: May 2005

AbstractAsparagine‐linked glycans of bovine fetuin were separated by capillary gel electrophoresis after enzymatic release (peptide‐N‐glycosidase F) and labeling via reductive amination by a fluorescent dye, 1‐aminopyrene‐3,6‐8‐trisulfonate (APTS). At low separation pH (2.5) only two dominant peaks were observed. Increasing the separation buffer pH to 4.75 resulted in complete separation of two primary doublets and several minor peaks from the fetuin N‐linked glycan pool. Two of the four major peaks were spiked with purified individual standards and were identified as trisialylated triantennary structures with different sialylation linkages. The other two larger peaks were postulated to be tetrasialylated triantennary structures, based on calculations considering their corresponding glucose unit (GU) values. Effects of the electrophoretic separation parameters, such as gel concentration, electric field strength and temperature on the migration behavior of the two major doublets of the fetuin glycan pool were also thoroughly examined. Our data suggest that the capillary gel electrophoresis separation of the multisialylated branched oligosaccharides with different linkage isomers, released from bovine fetuin, is fundamentally based on their degree of sialylation and hydrodynamic volumes.