bluuebunny/crossref_metadata_2025_split huggingface dataset:
Steady‐state heat transfer and thermal zone spreading in gel isoelectric focusing
Authors: Righetti Pier Giorgio , Bello Michael S. 
Published: May 2005

AbstractThe zone spreading caused by a transverse pH profile due to a temperature gradient through the thickness of a gel slab is estimated. The temperature difference (ΔT) between the upper and lower gel surfaces can be calculated as a function of the electric power dissipated in the gel and the gel dimensions. It is found that when ΔTis only 1 °C the zone spreading due to this thermal excursion is as high as 0.5 mm. Thus, an admissible ΔTis found to be equal to 0.2°C, since this corresponds to a thermal zone spreading of only 0.1 mm,i.e.the same order of magnitude as the spatial resolution of a laser scanner. A thermal zone spreading of 0.1 mm is compatible with a resolving power of 0.01 pH unit, the current limit of conventional isoelectric focusing in amphoteric buffers. A requirement for the thickness of a gel slab is formulated:e.g., at 40 W applied power (over a gel surface area of 25×11 cm), a thermal zone spreading limited to 0.1 mm can only be obtained with a gel thickness of approximately 170 μm.
Temperature gradient gel electrophoresis: Rapid detection of alpha‐1‐antitrypsin deficiency carriers
Authors: Hinney Anke , Luckenbach Christine , Ritter Horst 
Published: May 2005

AbstractThe homozygous state of the alpha‐1‐antitrypsin (αAT) deficiency variant Z is associated with severe liver damage in early childhood and progressive lung emphysema in adulthood. A single base transition (G to A in codon 342) in exon V is causing the severe disease. The Glu342 to Lys342 mutation can be detected conventionally by isoelectric focusing (IEF) or on the DNA level by the newly developed method of temperature gradient gel electrophoresis (TGGE). It is the aim of this study to describe the TGGE technique, to compare the results with conventional IEF, and to discuss its efficiency for different diagnostic applications.
A rapid and efficient method for the screening of acid phosphatase 1 in young tomato seedlings, and for the identification of root‐knot nematode species using miniaturized polyacrylamide gel electrophoresis
Authors: Cap Guillermo B. , Roberts Philip A. 
Published: May 2005

AbstractA relatively rapid and highly sensitive miniaturized polyacrylamide gel electrophoresis technique is described for the analysis of certain isozymes from single cotyledons of tomato seedlings and from single females of the root‐knot nematode (Meloidogyne spp.). Homogenates from single tomato cotyledons (7, 14, 21, and 28 days old) were electrophoresed and stained for acid phosphatase 1 (Aps 1) activity. Cotyledons from plants of all the above age groups showed good Aps 1 activity. Nondestructive screening for tomato Aps 1 is therefore feasible, using very small samples, from as young as 7‐day‐old tomato seedlings. This could be of important use in expediting root‐knot nematode resistance (based on the Aps 1‐linked resistance gene Mi) screening for breeding programs, or F1 testing for seed production purposes. In addition, the mini‐polyacrylamide gel electrophoresis technique was useful for determination of the Aps 1 allelic contribution to the total enzyme activity. The system was also used to detect malate dehydrogenase and esterase isozyme activity from single adult females of the four common root‐knot nematodes, Meloidogyne arenaria, M. hapla, M. incognita, and M. javanica, with equally good results, enabling species discrimination.
Increased electrophoretic mobility of sodium sulfitetreated jack bean urease
Authors: de Llano J. Javier Martín , Gavilanes José G. 
Published: May 2005

AbstractSodium sulfite is a widely used activity‐protective agent for the storage of urease. However, this reagent produces a 10% increase in the anodic electrophoretic mobility of native urease. Changes in the hydrodynamic properties of the enzyme are not involved in that modification. The observed change is related to an increased negative charge of the protein molecule in the presence of sodium sulfite. The results are discussed in terms of sulfitolysis of the single disulfide bond in the urease monomer. It is remarkable that the modification occurs at neutral pH. Our results show that removing sodium sulfite and reversing its effect by treatment with 2‐mercaptoethanol are required prior to any study involving native urease.
Two‐dimensional electrophoretic analysis of transformation‐sensitive polypeptides during chemically, spontaneously, and oncogene‐induced transformation of rat liver epithelial cells
Authors: Wirth Peter J. , Luo Lin‐di , Fujimoto Yoshinori , Bisgaard Hanne C. 
Published: May 2005

AbstractRecently, we described the establishment of a computerized database of rat liver epithelial (RLE) cellular polypeptides (Wirth et al., Electrophoresis, 1991, 12, 931–954). This database has now been expanded to include the analysis of cellular polypeptide alterations during chemically (aflatoxin B1; AFB), spontaneously, and oncogene (v‐Ha‐ras, v‐raf, and v‐myc/v‐raf)‐induced transformation of RLE cells. Two‐dimensional mapping of [35S]methionine‐labeled whole cell lysate, cell‐free in vitro translation products and [32P]orthophosphate‐labeled polypeptides revealed subsets of polypeptides specific for each transformation modality. A search of the RLE protein database indicated the specific subcellular location for the majority of these transformation‐sensitive proteins. Significant alterations in the expression of the extracellular matrix protein, fibronectin, as well as tropomyosin‐ and intermediate filament‐related polypeptides (vimentin, ß‐tubulin, the cytokeratins, and actin) were observed among the various transformant cell lines. Immunoprecipitation and Western immunoblot analysis of tropomyosin expression in four individual AFB‐, as well as four spontaneously induced, and each of the oncogene‐transformed cell lines indicated that five major tropomyosin (Tm 1–5) isoforms were variably expressed in the various cell lines, including one polypeptide tentatively identified as Tm6. Whereas alterations in tropomyosin expression appeared to be transformation‐specific, alterations in the individual intermediate filament polypeptides were related more to the differentiation state of the individual cell lines rather than to the transformation phenotype. These studies extend our earlier efforts toward the establishment of a comprehensive computerized database of RLE cellular proteins and demonstrates how such a database may serve as a useful source for studies concerning the regulation of growth and differentiation as well as transformation of RLE cells.
DNA‐protein interaction analysis using polyacrylamide gel electrophoresis and a simple and sensitive UV crosslinking procedure
Authors: Dooley Steven , Welter Cornelius , Blin Nikolaus 
Published: May 2005

AbstractA simple and reproducible technique for DNA‐protein interaction analysis is described using UV crosslinking and polyacrylamide gel electrophoresis, leading to visualization of the complexes as distinct and strong signals. It avoids incorporation of bromodeoxyuridine (BrdU) into DNA and requires no special equipment. It was successfully applied to an oligonucleotide sequence from within the first intron of the mouse myb proto‐oncogene and nuclear extracts from a myb‐expressing cell line.
Inter‐α‐trypsin inhibitor polymorphism in African blacks
Authors: Vogt Ulrike , Gürtler Lutz , Cleve Hartwig 
Published: May 2005

AbstractThe inter‐α‐trypsin inhibitor (ITI) polymorphism was analyzed in an African Negroid population using polyacrylamide gel isoelectic focusing and subsequent immunoblotting. Gene frequencies of ITI*1, ITI*2, ITI*3 and ITI*4 were calculated to be 0.564, 0.083, 0.337 and 0.004, respectively. One unknown rare allele, ITI*6 determines further phenotypes in combination with the alleles ITI*1 and ITI*3 Gene frequency of ITI*6 was calculated to be 0.012. The common alleles are represented by ITI*1 and ITI*3. The allele distribution is therefore different from European and Asian populations.
Deletion studies to reveal the basis for size discrepancy in proliferating cell nuclear antigen
Authors: Liang Chien‐Ping , Lee Yeu‐Chin , Liu Yin‐Chang 
Published: May 2005

AbstractProliferating cell nuclear antigen (PCNA), an essential component for DNA replication in eukaryotes, is a highly conserved nonhistone nuclear protein of 261 amino acids. The molecular weight of mammalian PCNA, estimated by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE), differs notably from that predicted by the cDNA sequences, that is, 36000 in comparison with 29261 and 28748 for human and rat PCNA, respectively. To investigate if this discrepancy is due to posttranslational modifications, we studied the PCNA protein synthesized by an in vitro transcription/translation system as well as the protein overproduced in bacteria. We found that both PCNA protein samples were indistinguishable from the authentic protein from the protein mobility in SDS‐PAGE. The finding indicates that the size discrepancy is not due to the posttranslational modifications. Hence, the size discrepancy may be due to the protein sequence per se, namely a sequence‐related anomaly in SDS‐PAGE. Results from the analyses of a series of PCNA derivatives with various lengths of C‐ or N‐terminal deletion indicate that the putative sequence is in the region of residues 128–150.
Improvement of BF typing and of BF F subtyping after neuraminidase treatment in the unconverted and converted factor B
Authors: Siemens Ingbert , Geserick Gunther , Mauff Gottfried , Correns Andreas , Schröder Helga 
Published: May 2005

AbstractWhen neuraminidase‐treated sera are analyzed by agarose gel isoelectric focusing, the factor B (BF) banding pattern is reduced to predominantly one major band without cathodically positioned bands. This not only makes unequivocal typing of BF allotypes possible but also the reliable distinction of all BF F subtype phenotypes with delimitation of “BF F subtype variants”. With this new method, serum aging affects the BF determination to a lesser extent than when applying methods that separate native sera. We show that sialylation is not responsible for the BF F subtype polymorphism. All of the investigated BF allotype bands, including those characteristic of the subtypes, show functional hemolytic activity. The banding pattern after removal of neuraminic acid residues ranges from pH 6.8 to 7.3 for factor B, from pH 5.3 to 5.9 for the Ba fragment, and from pH 8.2 to 8.7 for the Bb fragment. The protein structure of factor B is also discussed. Eliminating the superimposition of bands in different BF allotypes, as demonstrated by these methods, proved to be necessary for the detection of hypomorphic BF gene products (BF QL), which are expressed by assumed BF*Q0 alleles in heterozygous genotypes. This allows investigation of BF*Q0 alleles on a protein level, which complements molecular genetic approaches.
v‐erbB oncogene expression accounts for most variations in protein synthesis after avian erythroblastosis virus infection of chicken embryo fibroblasts: A two‐dimensional electrophoresis study
Authors: Desbois Christelle , Gandrillon Olivier , Samarut Jacques , Madjar Jean‐Jacques 
Published: May 2005

AbstractThe effect of the v‐erbA and/or v‐erbB oncogenes on cellular gene expression was investigated after separation by two‐dimensional polyacrylamide gel electrophoresis of [35S]methionine‐labelled proteins from chicken embryo fibroblasts (CEF), infected by either the avian erythroblastosis virus (AEV) carrying both oncogenes, or by viruses carrying only one of them. We observed significant changes in the synthesis of 34 proteins in AEV‐transformed CEF as compared with control cells. The synthesis of 24 of them was increased while the synthesis of the other 10 proteins was decreased. The expression of v‐erbB alone is necessary and sufficient to induce changes in the synthesis of 27 proteins while the 7 remaining modifications are observed only in cells expressing v‐erbB together with v‐erbA. Moreover, the deregulation of protein synthesis by v‐erbB‐expressing viruses was correlated with the morphological transformation state of cells.
A new concept for sequencing DNA by capillary electrophoresis
Authors: Noolandi Jaan 
Published: May 2005

AbstractIt is proposed that the scaling symmetry of constant charge density with increasing molecular weight, which prevents the separation by electrophoresis of DNA molecules in solution (with respect to molecular weight) be broken by the attachment of a perturbing entity (protein, virus or charged sphere) to one end of the molecule. An application of this idea to a concept for sequencing DNA by capillary electrophoresis is discussed, and the possibility of using the reattachment of the RecA protein to separate large segments of DNA in solution by electrophoresis following sequence‐specific cleavage is mentioned.
Separation of proteins by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis in the presence of soluble, aqueous polymers: Ficoll and polyvinylpyrrolidone
Authors: Gersten Douglas M. , Bijwaard Karen E. 
Published: May 2005

AbstractIn previous studies we have demonstrated that water soluble polymers of dextran and methylcellulose, when incorporated into sodium dodecyl sulfate (SDS)‐polyacrylamide gels, can be used to sharpen the protein bands. We have extended these studies to enhance the separations in the mid‐molecular weight range. Enhanced separation of protein molecular weight markers between 30000–67000 was best achieved in SDS‐gels containing 2.5% polyvinylpyrrolidone (average molecular weight 10000).
Classification of crossed immunoelectrophoretic patterns using digital image processing and artificial neural networks
Authors: Søndergaard Ib , Krath Britta Naimi , Hagerup Martin 
Published: May 2005

AbstractA method is presented which makes it possible to present crossed immunoelectrophoretic patterns to an artificial neural network. The electrophoretic patterns are presented for the artificial neural network as three‐dimensional vectors and it is shown that it is possible with this representation to train the network to learn the patterns and classify them. It was found that the ability to generalize was substantially increased by the addition of noise to the input patterns during training. Furthermore, the addition of noise decreased the number of presentations needed to reach the predetermined error level. The trained neural network was able to classify all distorted patterns correctly within an error range of 1%.
Sex identification by polymerase chain reaction of α‐satellite in aged tissue samples
Authors: Fattorini Paolo , Cacció Simone , Gustincih Stefano , Florian Fiorella , Altamura Bruno M. , Graziosi Giorgio 
Published: May 2005

AbstractIn vitroamplification of the alphoid repeated sequences clustered in the centromeric regions of the human × and Y chromosomes was carried out. A modification of the amplification conditions reported by Witt and Erickson [1] gave clear amplification bands from 10 pg of genomic DNA template. Results were obtained from the following aged tissue samples: 1 μL three‐year‐old blood stain, one‐year‐old single hair roots, six‐month‐old saliva stains and twelve‐year‐old bone fragments. Good results were obtained even when the template was artificially fragmented into fragments of less than 1 kbp.
Gel electrophoresis of giant proteins: Solubilization and silver‐staining of titin and nebulin from single muscle fiber segments
Authors: Granzier Henk L. M. , Wang Kuan 
Published: May 2005

AbstractGiant proteins in the megadalton range (> 0.5 MDa) appear to play important structural and functional roles in striated muscle. Titin (∼ 3 MDa) is involved in the generation of resting tension and the assembly and stability of the sarcomere in skeletal and cardiac muscle tissues, while nebulin (∼ 0.7 MDa) is thought to regulate thin filament length in skeletal muscle. Sodium dodecyl sulfate (SDS)‐gel electrophoresis is an important tool in revealing the size, quantity and integrity of these giant proteins in muscle tissues. We report here a method for solubilizing, detecting and quantifying titin and nebulin from short segments of single fibers of the rabbit psoas muscle. Muscle proteins ranging from 15 kDa to 3MDa were resolved on 3.3–12% gradient polyacrylamide gels that were silver‐stained and quantitated by densitometry. Presoaking fiber segments in a low ionic strength pH 8.4 buffer enhances the amount of solubilized titin and nebulin. Solubilizing the presoaked fiber segments with SDS at 60°C for 60 s maximizes the amount of intact titin; solubilizing at higher temperatures causes extensive degradation of titin. Detection sensitivity is sufficient to study titin and nebulin in fiber segments as short as 120 μm.
Extracellular protein secretion of cultured normal and Pagetic osteoblasts
Authors: Hankey David P. , Hughes Anne E. , Mollan Raymond A. B. , Nicholas Richard M. 
Published: May 2005

AbstractTwo‐dimensional polyacrylamide gel electrophoresis (2‐D PAGE) has been used to map the extracellular secretory activity of normal osteoblasts. The proteins osteonectin, bone sialoprotein, and both the C‐telopeptide of collagen I together with collagen I have now been positionally identified. In addition the secretory differences which exist between normal and Pagetic osteoblasts have been mapped with the Pagetic osteoblasts shown to consistently secrete an altered 30 kDa C‐telopeptide of collagen type I. The use of the diphosphonate Pamidronate in the treatment of Paget's disease of bone has beneficial effects with suppression of the bone isoenzyme marker alkaline phosphatase. It has been reported that diphosphonates directly inhibit human osteoblast secretory function as well as osteoclast metabolism. The effects of Pamidronate on the secretory activity of normal and Pagetic osteoblast cultures was also investigated. The extracellular protein secretion of normal and Pagetic osteoblasts was not affected by Pamidronate treatment as assessed by 2‐D PAGE. This technique allows a comprehensive multiparameter assessment of extracellular secretory activity in normal and diseased states. The findings show that the Pagetic osteoblasts cultured in vitro are functionally abnormal and they support the hypothesis that the underlying problem in Paget's disease is characterised by disorder of osteoblast and osteoclast interactions.
Rapid detection of mitochondrial DNA polymorphism by photoactivatable biotin
Authors: Tsuchida Shuichi , Ikemoto Shigenori 
Published: May 2005

AbstractA new method for the detection of mitochondrial (mt)DNA polymorphism by agarose gel electrophoresis is presented. In this method, after mtDNA digestions with endonucleases, the mtDNA fragments are labeled directly with photoactivatable biotin in the digestion buffer, separated by agarose gel electrophoresis, transferred to a nylon membrane and detected enzymatically using streptavidinalkaline phosphatase, Nitroblue tetrazolium (NBT) and 5‐bromo‐4‐chloro‐3‐indolylphosphate (BCIP). This method is useful for the detection of mtDNA digestion patterns by agarose gel electrophoresis, especially when enough purified mtDNA cannot be obtained and the radioisotope cannot be handled because of several restrictions.
Detection of DNA damage‐recognition proteins using the band‐shift assay and Southwestern hybridization
Authors: Protić Miroslava , Levine Arthur S. 
Published: May 2005

AbstractWe describe electrophoresis and biochemical conditions that allow detection of damaged DNA‐binding proteins in cell extracts. In addition, we present an overview of the damage‐recognition DNA‐binding proteins from eukaryotic cells and discuss their hypothetical role in DNA repair.
Electrophoretic analysis of protein‐induced DNA bending and twist changes
Authors: Niederweis Michael , Hillen Wolfgang 
Published: May 2005

AbstractDNA fragments with a varied phasing of two intrinsic bends at their ends and a tet operator in‐between were constructed to determine Tet repressor‐induced twist changes in the operator DNA. These distance variants show a sinusoidal dependence of their electrophoretic mobilities on the phasing of their bends in polyacrylamide gels. Complex formation with Tet repressor leads to a displacement of the first minimum, indicating an unwinding of the tet operator DNA. Model calculations were performed to assess the contribution of Tet repressor‐induced DNA bending to this result. They revealed that the amplitude of the electrophoretic mobilities of the distance variants may be used as a parameter to separate the effects of Tet repressor‐induced twist change and bending. The systematic analysis presented here may help to improve the quantitative interpretation of gel shifts and promote the use of this highly sensitive method to gain structural information about protein‐DNA complexes.
Automated Ferguson analysis of glycoproteins by capillary electrophoresis using a replaceable sieving matrix
Authors: Werner William E. , Demorest David M. , Wiktorowicz John E. 
Published: May 2005

AbstractObtaining accurate molecular weight estimates for glycoproteins by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE) has been difficult due to the lack of SDS binding by the carbohydrate moieties of the proteins. This leads to lower charge‐to‐mass ratios for SDS‐glycoprotein complexes, resulting in over‐estimation of molecular weights by SDS‐PAGE. In order to minimize these inaccuracies for proteins with abnormal charge‐to‐mass ratios, a Ferguson plot may be employed. This application requires the determination of relative mobilities for standard proteins in addition to unknowns at several different gel concentrations. Historically, this technique has not been popular because it requires time‐consuming preparation of gels with varying matrix concentrations, electrophoresis, and staining/destaining of gels. In this paper a procedure is demonstrated which automatically generates all of the data required for a Ferguson plot using a replaceable sieving matrix (thereby eliminating gel polymerization) in a capillary format. In addition, this technique possesses the advantages inherent to capillary electrophoresis, namely, very fast separation times, and on‐line monitoring which allows quantitation and precludes post‐separation staining and destaining of gels.