bluuebunny/crossref_metadata_2025_split huggingface dataset:
Biomedical and clinical applications of automated single cell electrophoresis
Authors: Schütt Wolfgang , Thomaneck Uwe , Knippel Eberhard , Rychly Joachim , Klinkmann Horst 
Published: May 2005

AbstractThe automated single cell electrophoresis microscope Parmoquant has been used for the discrimination of lymphocytes and the study of the interaction of substances with cells and synthetic particles. Electrophoretic histograms allowed the determination of changes in the proportion of lymphocyte populations after kidney transplantation, during dialysis treatment, open heart surgery and during pregnancy. Discrimination of leukemic cells on the basis of electrophoresis was used as an additional parameter in diagnosis. In a mouse tumor model, histogram determination enabled the in vivo effect of tumor necrosis factor on immune cells to be evaluted. Cell electrophoresis was shown to be suitable to detect the influence of antibodies, lectins and bacteria on the cell surface. Protein adsorption was studied on synthetic particles using cell electrophoresis. This method was applied to investigate the phenomena of blood interaction with biomaterials used in artifical organs and to determine differences in the protein composition of serum or other body fluids connected with diseases.
Separation of rat pituitary secretory granules by continuous flow electrophoresis
Authors: Hayes Daniel , Exton Carrie , Salada Thomas , Shellenberger Kathy , Waddle Jenny , Hymer W. C. 
Published: May 2005

AbstractThe separation of growth hormone‐containing cytoplasmic secretory granules from the rat pituitary gland by continuous flow electrophoresis is described. The results are consistent with the hypothesis that granule subpopulations can be separated due to differences in surface charge; these, in turn, may be related to the oligomeric state of the hormone.
Preparative separation of pyrogens from proteins by isoelectric focusing using a multicompartment electrolyser with Immobiline membranes
Authors: Lucas J. , Faupel M. , Goecking C. 
Published: May 2005

AbstractDue to the nature of lipopolysaccharide endotoxin structures of bacterial pyrogens, their removal from solutions containing therapeutic proteins is often a problem in the pharmaceutical industry. In this report we describe the application of electromotive force to dislodge lipopolysaccharide endotoxins from proteins. This was performed by employing a multicompartment electrolyzer fitted with Immobiline membranes of specified pIs. A thousand‐fold reduction of endotoxin could be achieved in the model test system described. This contribution describes the use of a new recycling isoelectric focusing approach without the use of carrier ampholytes.
Denaturing gradient gel electrophoretic analysis of minisatellite alleles
Authors: Uitterlinden André G. , Vijg Jan 
Published: May 2005

AbstractBy two‐dimensional DNA fingerprinting, an electrophoretic method which combines separation according to size with separation in a denaturing gradient, virtually all minisatellite sequences detected with a minisatellite core probe can be resolved (Uitterlinden et al., Proc. Natl. Acad. Sci. USA. 1989, 86, 2742–2746). To investigate the electrophoretic behavior in denaturing gradient gels of allelic restriction fragments containing minisatellite sequences, we analyzed alleles of the two highly polymorphic minisatellite loci D7S22 and D2S44. The results obtained indicate that for these loci, depending on the restriction enzyme used to digest genomic DNA, alleles of different sizes migrate to regions of similar denaturant concentration, i.e. to isothermal positions in the denaturing gradient. Denaturing gradient gel electrophoresis also allows for the discrimination of restriction fragments which are the result of the presence of internal recognition sites in the minisatellite and, therefore, to distinguish between VNTR and restriction site polymorphisms.
Detection of β‐glucanase activity on various β‐1,3 and β‐ 1,4‐glucans after native and denaturing polyacrylamide gel electrophoresis
Authors: Côté François , El Ouakfaoui Souad , Asselin Alain 
Published: May 2005

Abstractβ‐Glucanases were detected after polyacrylamide gel electrophoresis under native and denaturing conditions using various β‐1, 3‐ and β‐1, 4‐glucans, including mixed glucans (laminarin, pachyman, carboxymethyl cellulose, lichenan and barley β‐glucan). After electrophoresis and incubation of gels, substrates incorporated into polyacrylamide gels were stained with specific fluorochromes, Sirofluor for β‐1, 3 linkages and Calcofluor White M2R for β‐1, 4 linkages. Under UV illumination, lysis zones appeared as dark bands against a fluorescent background. Enzymes of bacterial, fungal and plant sources could be revealed sequentially in gles containing mixed β‐(1.3) (1,4)‐glucans by staining first with sirofluor followed by staining with Calcofluor White M2R. Active profiles were more diverse when substrates were stained with sirofluor. The use of purified sirofluor at pH 11.5 compared with Aniline Blue at pH 8.6 allowed better detection of β‐1, 3‐glucanase activities. In gels containing laminarin stained with sirofluor, bands exhibiting a more intense fluorescence than the background fluorescence were observed in addition to dark nonfluorescent bands. It is postulated that these two types of β‐1, 3‐glucanase activities differ by their enzymatic action (partial versus extensive hydrolysis). Analysis of fungal extracts using denaturing gels embedded with various β‐glucans displayed lysis bands migrating between 32 and 35 kDa.
Cationic detergent in agarose for improved electrophoresis of cationic proteins
Authors: Reimert Claus M. , Bøg‐Hansen Thorkild C. 
Published: May 2005

AbstractInclusion of the cationic detergent N‐cetyl‐N,N,N‐trimethylammonium bromide (CTAB) in normal agarose gels increases the electrophoretic separation and banding pattern of cationic proteins. This is shown for an extract of cationic granular proteins from polymorphonuclear granulocytes.
Oligonucleotide fingerprinting with (CAC)5: Nonradioactive in‐gel hybridization and isolation of individual hypervariable loci
Authors: Zischler Hans , Hinkkanen Ari , Studer Roland 
Published: May 2005

AbstractThe first topic to be treated in this paper is the nonradioactive DNA fingerprinting by means of in‐gel hybridization with digoxigenated (CAC)5. Besides the fact that timeconsuming Southern blotting can be avoided, the dried agarose is an excellent matrix to produce background‐free nonradioactive DNA fingerprints. There is no tendency of either the oligonucleotide probe or the antibody towards unspecific binding to the dried agarose. Prehybridization and blocking steps are therefore superfluous. Furthermore, we will discuss what effect the degree of crosslinking of the antibodyenzyme conjugates has. The second topic concerns the isolation and characterization of locus‐specific probes from a human (CAC)5 fingerprint. The isolation and characterization of one variable probe, by screening complete genomic libraries, is described and discussed. This probe is compared to a hypervariable single‐copy probe, isolated from a size‐enriched genomic library. The sequence of the repeat flanking locus‐specific probe is presented and a semi‐specific, adaptor‐mediated polymerase chain reaction was designed to amplify (CAC)n/(GTG)n flanking sequences.
Databases, quality control and interpretation of DNA profiling in the Home Office Forensic Science Service
Authors: Gill Peter , Evett Ian W. , Woodroffe Susan , Lygo Joan E. , Millican Emma , Webster Mark 
Published: May 2005

AbstractThe history of DNA profiling in the Home Office Forensic Science Service began with the introduction of multilocus probes into casework in 1986. The use of singlelocus probes was introduced in 1990, supported by databases of three ethnic groups; interpretation is backed up using a Bayesian approach. Databases were compiled using an image analysis computing system. Quality control systems are described, detailing requirements before a sample can be included in the database.
A discussion of the robustness of methods for assessing the evidential value of DNA single locus profiles in crime investigations
Authors: Evett Ian W. , Gill Peter 
Published: May 2005

AbstractWe discuss three issues relating to the evaluation of DNA single locus probe comparisons in forensic casework: band shift, database size, and population stratification. Illustrations are from experiments carried out in the Forensic Science Service as part of our preparations to implement the technique.
Secretion of lipid‐poor nascent human apolopoprotein apoAI, apoCIII, and apoE by cell clones expressing the corresponding genes
Authors: Hussain M. Mahmood , Roghani Ali , Cladaras Christos , Zanni Eleni E. , Zannis Vassilis I. 
Published: May 2005

AbstractThe human apolipoprotein apoAI, apoCIII, and apoE genes were placed under the control of the mouse metallothionein 1 promoter in a bovine papilloma virus vector that also contained the human metallothionein IA gene. Following transfection of mouse C127 cells with the expression vector, cell clones resistant to Cd2+ were selected and found to express in high abundance specific apolipoprotein genes. Individual cell clones expressing apoAI, apoCIII, or apoE genes were used further to study the isoprotein composition and the flotation properties of the corresponding nascent apolipoproteins. It was found that the lipoproteins secreted by cell clones ex‐pressing the apoAL, apoCIII, and apoE genes consisted of the proapoAI disialylated form of apoCIII (apoCIIIs2) and mainly sialylated forms of apoE. Separation of the secreted apolipoproteins by density gradient ultracentrifugation resulted in limited flotation of nascent apoAI, apoE and apoCIII in the high density lipoprotein (HDL) fraction. Similar analysis in the presence of human serum increased the flotation of apoAI, apoE, and apoCIII to 6.5‐, 4.5‐, and 5.5‐fold, respectively, and resulted in their redistribution to various lipoprotein fractions. HDL increased the flotation of apoAI to 12‐fold and very low density lipoprotein (VLDL) increased the flotation of apoCIII and apoE to 6.5‐ and 5.5‐fold, respectively. These findings suggest that in the cell system used, the majority of nascent apoAI, apoCIII and apoE is secreted in the lipid‐poor form, which then associates extracellularly with preexisting lipoproteins.
A reliable two‐dimensional gel electrophoresis procedure for separating neural proteins
Authors: Semple‐Rowland Susan L. , Adamus Grazyna , Cohen Robert J. , Ulshafer Robert J. 
Published: May 2005

AbstractA facile two‐dimensional gel electrophoresis procedure has been developed for the analysis of neural tissue proteins which eliminates the serious problems associated with protein insolubility at the point of sample application onto polymerized first dimension isoelectric focusing gels. This was accomplished by combining the methods of two previously published procedures. Our proecedure provides an alternative method to the complex gel systems often employed for less soluble proteins, and yields very reproducible, high resolution separations. This procedure, which is in routing use in our laboratories for the analysis of total proteins extracted from retina and brain, produces protein patterns that are easily compared using both visual and computer‐assisted image analysis techniques. Presented here are the results of a set of experiments designed to identify proteins unique to retina. This procedure should be useful to investigators studying protein changes resulting from genetic mutation, development, drug treatment or disease, in neural tissue as well as in virtually all other tissues.
Barley cultivar discrimination: I. Sodium dodecyl sulfate‐polyacrylamide gel electrophoresis and glycoprotein blotting
Authors: Weiss Walter , Postel Wilhelm , Görg Angelika 
Published: May 2005

AbstractTwo different methods of detecting electroblotted glycoproteins after sodium dodecyl sulfate‐polyacrylamide gel electrophoresis of Tris‐buffer soluble barley seed proteins were examined for their applicability for barley cultivar discrimination. These are the highly specific, lectin‐based concanavalin A/peroxidase method and the more general periodate/danyslhydrazine method. The results of the periodate/dansylhydrazine method enabled us to divide the 20 examined cultivars into three groups, whereas the more sensitive concanavalin A/peroxidase method revealed six different glycoprotein patterns. In comparison, sodium dodecyl sulfate‐polyacrylamide gel electrophoresis and silver staining of the alcoholsoluble barley seed proteins (hordeins) gave nine different banding patterns. A combination of hordein electrophoresis together with glycoprotein staining by the concanavalin A/peroxidase method made it possible to classify the cultivars into twelve groups, the largest of which contained four cultivars. The qualitative expression of the glycoprotein patterns seemed to be independent of growth conditions, whereas the band intensities obviously were not. As a whole, glycoprotein blotting is a valuable supplement to sodium dodecyl sulfate‐polyacrylamide gel electrophoresis of hordeins in barley cultivar discrimination.
Gel electrophoresis of polystyrene particles in glutaraldehyde crosslinked polyvinyl alcohol
Authors: Pospichal Jan , Tietz Dietmar , Ittyerah T. Roy , Halpern David , Chrambach Andreas 
Published: May 2005

AbstractPolystyrene sulfate and carboxylate particles (19–189 m radius) were subjected to electrophoresis in glutaraldehyde crosslinked polyvinyl alcohol of molecular weight 25.000 and 650.000 Da at various concentrations. The degree of crosslinking is severely limited by the mechanical properties of the gels that deteriorate beyond a glutaraldehyde concentration which decreases with increasing polyvinyl alcohol chain length. The effective fiber radius of the short‐chain and long‐chain polymer fiber was 45 ± 25 and 131 ± 47 nm, respectively. Thus, these media do not significantly exceed the apparent fiber thicknes of agarose, are more difficult to prepare – but are well‐defined synthetic product rather than natural ones, and have the advantage of carrying no net charge and can therefore be expected to exhibit no electroendosmosis.
Observation of orientation and relaxation of protein‐sodium dodecyl sulfate complexes during pulsed intermittent field polyacrylamide gel electrophoresis
Authors: Brassard Éric , Turmel Chantal , Noolandi Jaan 
Published: May 2005

AbstractPolyacrylamide gel electrophoresis (PAGE) of proteins denatured with SDS (sodium dodecyl sulfate) has been used successfully to separate proteins according to their molecular mass. In spite of the extensive use of this technique, the motion of the protein‐SDS complex in a polyacrylamide gel is still not understood. Here we report on the observation of the orientation (in the field direction) and relaxation of protein‐SDS complexes during pulsed intermittent field PAGE experiments. The results give an indication of the stiffness of the molecules and may be useful for the development of a technique to improve the separation of large proteins using pulsed electric fields.
A miniaturized electrode configuration for isoelectric focusing
Authors: Megow Dirk , Jacobasch Gisela 
Published: May 2005

AbstractAn electrode configuration is described which allows fast isoelectric focusing (IEF) with conventional IEF systems. The equipment, which can be fixed on the cooling plate of a conventional IEF system, consists of a base plate on which flappable electrode holders are fastened. The handling is simple and needs only little time. Graphite rods are used as electrodes, thus avoiding the use of buffer strips. Samples are applied with special applicator strips – permitting the analysis of up to 19 samples on a 50 × 40 mm polyacrylamide gel and up to 44 samples on a 100 × 70 mm gel. Only 30 min are needed for one IEF run.
A novel method for specific visualisation of serum albumin in polyacrylamide gels by iodine staining
Authors: Lee Kuo‐Kau , Ellis Anthony E. 
Published: May 2005

AbstractBovine serum, bovine serum albumins (delipidated or globulin free or Fraction V), rabbit serum, rabbit serum albumin, Atlantic salmon serum, purified Atlantic salmon serum albumin, human plasma α2‐macroglobulin, hemocyanin, trypsin inhibitor, bovine transferrin and bovine lactoferrin were examined by a novel method for specific visualisation of albumins. In native polyacrylamide gel electrophoresis the albumins were visualised by iodine staining as a transparent spot against a brown background whilst the other proteins could not be visualised. It is suggested that the brown background was due to penetration of the gel by iodine while the chemical binding of iodine by albumin produced a decolourisation reaction. This novel method provides a fast and simple approach to identifying serum albumin in polyacrylamide gels.
A rapid (< 10 minute) electrophoresis method for identification of wheat varieties
Authors: Wrigley Colin W. , Gore Peter J. , Manusu H. Perry 
Published: May 2005

AbstractConventional procedures for electrophoretic identification of gain samples according to variety are too slow to permit checking at the time of delivery. The method described permits electrophoretic identification within an hour. It involves extraction of gliadin proteins from crushed grain with 6% urea solution or ethylene glycol, cathodic electrophoresis for 9 min at 300 V in a Micrograd gel (MG 315 from Gradipore Ltd, Sydney, Australia) using sodium lactate buffer (pH 3.1), and staining in Gradipore (at about 50°C). Distinction between a set of Australian varieties was similar to that obtainable with the Australian Standard Procedure.
Counterflow affinity isotachophoresis on cellulose acetate membranes
Authors: Schranz Dorota , Morkowski Stanislaw , Karamova Eleonora R. , Abelev Garri I. 
Published: May 2005

AbstractCounterflow isotachophoresis on cellulose acetate membranes of human alphafetoprntein (AFP) was performed with concanavalin A, lentil lectin, and castor bean lectin driven by electroendosmotic counterflow. This counterflow caused a uniform stream of lectin to migrate towards the cathode against AFP with carrier ampholytes in steady‐state position. Retardation of microheterogeneity forms bound to lectins was observed, giving results comparable to standard crossed affinity immunoelectrophoresis. Smaller amounts of lectins and more diluted samples of AFP could be used in the described method.
Analysis of the interaction between human plasma fibronectin and gelatin by affinity electrophoresis
Authors: Kashiwagi Shiro , Nakamura Kazuyuki , Takeo Kazusuke , Takasago Teiichi , Uchimichi Akihiko , Ito Haruhide 
Published: May 2005

AbstractThe interaction between human plasma fibronectin and gelatin was analyzed by affinity electrophoresis, in which the fibronectin was subjected to electrophoresis in a 4% polyacrylamide gel in the presence and absence of gelatin, as an affinity ligand, and the fibronectin band was stained by an immunoblotting method. The apparent dissociation constants (Kd) of fibronectin for gelatin were calculated from affinity plots based on the original affinity equation at different pHs, urea concentrations, and temperatures. The fibronectin exhibited much lower affinity in the presence of urea. The Kds at 37°C were 1.49 × 10−7 M, 2.50 × 10−6 M, and 3.58 × 10−6 M with 2 M, 3 M, and 4 M urea, respectively. The van't Hoff plots of Kd values against absolute temperature (T) showed that the value of log Kd decreased in proportion to the increase in the value of 1/T within the range of 15–50°C. The standard enthalpy, the standard free energy change at 37°C, and the entropy change at 37°C for association were calculated to be −124.7 kJ/mol, −33.23 kJ/mol, and −295.1 J/mol/deg, respectively. These results suggest that a hydrophilic interaction, such as hydrogen bond or van der Waals interaction, plays an important role in the binding of plasma fibronectin to gelatin.
Demonstration of murine pancreas elastase and its interstrain variation by isoelectric focusing
Authors: von Deimling Otto H. , Gaa Andreas , Hameister Horst 
Published: May 2005

AbstractIsoelectric focusing between pH 9 and 11 was used for separation of murine pancreas proteases. One of these proteases is characterized by its preference for N‐acetyl‐L‐alanine‐α‐naphthylester as substrate and by its genetic linkage to bt, a coat color marker of chromosome 15. This protease was identified as elastase, and is probably elastase‐1 (ELA‐1). Because of the simple procedure and the excellent reproducibility of the focusing pattern, ELA‐1 is recommended as a useful marker for mouse chromosome 15.
Analysis of cerebrospinal fluid from patients with psychiatric and neurological disorders by two‐dimensional electrophoresis: Identification of disease‐associated polypeptides as fibrin fragments
Authors: Wildenauer Dieter B. , Körschenhausen Dirk , Hoechtlen Wilma , Ackenheil Manfred , Kehl Maria , Lottspeich Friedrich 
Published: May 2005

AbstractTwo‐dimensional electrophoresis (2‐DE) of cerebrospinal fluid (CSF) samples–from 347 patients with various psychiatric and neurological disorders–and subsequent silver staining revealed two additional polypeptides (Mr 40000) in 49% of 111 schizophrenics, 46 % of 43 schizoaffective patients, 36 % of 41 patients with affective disorders, 43 % of 28 patients with multiple sclerosis, but not in 25 patients without neurological symptomatology, nor in 9 patients with Lues, and in only 2 of 25 patients with AIDS. The two polypeptides, as detected by 2‐DE, eluted after size exclusion chromatography in fractions containing proteins with Mr > 200000. After 2‐DE of CSF samples, enriched by gel chromatography, the polypeptides were immobilized by blotting onto glass‐fiber membranes and subjected to N‐terminal sequencing. Polypeptide A was identified as β‐chain remnant (β2), derived from plasmin cleavage of fibrin(ogen). After size exclusion chromatography, 2‐DE, and Western blotting, polypeptide A and B, as well as several other spots, reacted with fibrinogen antibodies, suggesting that the polypeptides are subunits of a fibrin degradation complex.
Two‐dimensional polyacrylamide gel electrophoresis of human lung cancer: Qualitative aspects of tissue preparation in relation to histopathology
Authors: Franzén Bo , Iwabuchi Hiroshi , Kato Harabumi , Lindholm Johan , Auer Gert 
Published: May 2005

AbstractIt is of great importance to identify new objective markers significant for the diagnosis, treatment, and prognosis of human cancers. Cytometric assessment of the DNA content has been shown to be one parameter which correlates excellently with the clinical course of patients with solid cancers of the breast, prostate, and thyroid. However, additional specific marker polypeptides are needed in order to further improve the diagnostic and prognostic sensitivity. Two‐dimensional polyacrylamide gel electrophoresis (2‐DE) is a unique tool in this field of cancer research, but serious problems concerning intersample variations in the 2‐DE pattern must be solved because quantitative variations of potential marker polypeptides are expected to occur at low levels. In this study we examined a modified preparation method for tumor tissues, focusing on the relation between histopathological properties and 2‐DE gel quality. We selected a group of human lung cancers sharing similar prognosis; nevertheless, the 2‐DE patterns showed significant intersample but low intrasample variations. It is concluded that histopathological features, such as a local homogeneity, and the amounts of connective tissue and serum proteins are critical factors for the successful preparation and high quality of 2‐DE‐gels.