bluuebunny/crossref_metadata_2025_split huggingface dataset:
Multivariate statistical comparison of autoantibody‐repertoires (Western blots) by discriminant analysis
Authors: Zimmermann Christoph W. , Grus Franz‐Hermann , Dux Reinhold 
Published: May 2005

AbstractA procedure for the quantification and comparison of complex (auto‐) antibody repertoires of many individuals is described. It is based on multivariate statistical analyses of densitometric data of individual staining patterns, in this case Western blots. Implementation of algorithms can be based on standard software and hardware components. This procedure allows (i) the quantitative assessment of the reproducibility and reliability of any electrophoretic procedure, such as for the production of the immunological substrate, (ii) the statistical comparison of any group of staining patterns, e.g., derived from patients with autoimmune diseases or normal controls, (iii) the identification of the bands that contribute most to the differences between such groups, and (iv) the determination whether an unknown individual sample belongs to a known group. A statistical analysis of autoantibody repertoires has important possible applications: it detects spontaneous or therapeutically induced changes of repertoires, it identifies regions of interest, it supports the isolation of relevant antigens, and it will also be helpful in the diagnosis of autoimmune diseases.
Highly sensitive chiral analysis in on‐line combined chiral and achiral media by capillary zone electrophoresis
Authors: Krásenský Simon , Fanali Salvatore , Křivánková Ludmila , Boček Petr 
Published: May 2005

AbstractA new approach is described for highly sensitive chiral analyses by capillary zone electrophoresis, based on using an on‐line combination of two capillaries filled with either chiral selective or achiral background electrolytes (BGE). Thus, the BGEs are selected in such a way that the first capillary provides optimum chiral selectivity and the second one optimum detection sensitivity. Direct chiral separations of enantiomers of mandelic, m‐methoxymandelic, 3‐phenyllactic and 3‐indolelactic acids served as a model example for testing the approach proposed. The analyses were performed in a BGE containing acetate as a coion and L‐OH‐proline or aspartame as a chiral selector. For high sensitive analyses, an arrangement containing on‐line combined chiral and achiral media were tested in one or two capillaries coupled via a bifurcation block. A detection limit as low as 10−18 moles was reached in the column‐coupling system when the direct chiral separation was performed in the first capillary, filled with 20 mM acetate buffer, pH 4.4, containing 8 mM Cu (II) and 16 mM aspartame (L‐aspartyl‐L‐phenylalanine methylester) and separated enantiomers were detected in the second capillary, filled with 20 mM acetate buffer, pH 3.1. The principle described is of general use in cases where the separation and detection of analytes in question require mutually different BGEs to reach the optimum selectivity and sensitivity, respectively.
High‐resolution sodium dodecyl sulfate‐polyacrylamide gel electrophoresis and immunochemical identification of the 2X and embryonic myosin heavy chains in complex mixtures of isomyosins
Authors: Rossini Katia , Rizzi Corrado , Sandri Marco , Bruson Adonella , Carraro Ugo 
Published: May 2005

AbstractIn mammals myosin heavy chains (MHC) are polypeptides with a molecular mass of about 200 kDa whose isoforms can be identified by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS‐PAGE) and immunochemistry. Electrophoretic analysis is the only method for quantitating MHC profiles in single myofibers and/or cryostat sections of biopsied muscle. We present a method for SDS‐PAGE of adult rat skeletal muscle which resolves MHC into four bands: 1, 2B, 2X, and 2A from the faster to the slower migrating band. Furthermore, embryonic MHC can be also resolved in a complex mixture of isomyosins, e.g. developing or regenerating muscles. The method does not involve preparation of gradient gels or electrophoresis at low temperature. Improved reproducibility is obtained by: (i) modification of the sample buffer; (ii) use of 7% polyacrylamide in the separating gel; (iii) control of pH of running buffer by recirculation or change of the buffer during the run; and (iv) a 24 h run. The procedure is compatible with Coomassie Brilliant Blue, silver and immunoblot staining. Resolution is sufficient to permit transblotting of separated MHC after SDS‐PAGE. The different isoforms are easily identified with monoclonal antibodies. The technique provides an improved method to separate MHC and quantitate MHC2X and MHCemb in complex mixtures of MHC from a few cryostat sections of normal and diseased muscle.
Species identification of cyst and root‐knot nematodes from potato by electrophoresis of individual females
Authors: Karssen Gerrit , Van Hoenselaar Tine , Verkerk‐Bakker Brigitte , Janssen Richard 
Published: May 2005

AbstractTo carry out rapid and reliable species identification of Globodera and Meloidogyne specimens, two electrophoretic methods were modified and adapted for an automated electrophoresis system (PhastSystem, Pharmacia). Proteins of individual Globodera cysts were identified using isoelectric focusing and a sensitive silver stain. Proteins of young single Meloidogyne females were separated using polyacrylamide gradient gels and stained for the isozymes esterase and malate dehydrogenase.
Two‐dimensional gel electrophoresis of nasal and bronchoalveolar lavage fluids after occupational exposure
Authors: Lindahl Mats , Ståhlbom Bengt , Tagesson Christer 
Published: May 2005

AbstractHuman nasal lavage fluids (NLFs), and bronchoalveolar lavage fluids (BALFs) were analyzed with two‐dimensional gel electrophoresis (2‐DE) and the protein patterns were evaluated with a computerized imaging 2‐DE system. With silver staining about 1000 spots were detected in 10 μg samples of NLF or BALF. Both BALF and NLF 2‐DE patterns showed similarities to a reference plasma pattern and about 25 plasma proteins were identified in NLF as well as in BALF. Comparison showed that the levels of albumin and transferrin appeared to be slightly higher in BALF than NLF, while the levels of IgA, IgG and haptoglobin β were higher in NLF than in BALF. In contrast to BALF and blood plasma, NLF contained large amounts of a cluster of acidic proteins (pI 4.5–5.5) with molecular masses of 15–30 kDa. Distinct alterations in the NLF 2‐DE patterns were found in a worker who developed an asthmatic condition with bronchial hyperreactivity after exposure to organic acid anhydrides. After exposure, 14 protein spots were increased and one decreased by a factor of > 3 as compared to the levels before exposure and compared to healthy individuals. This is the first study indicating that 2‐DE of NLF may be used to investigate early changes in airway protein patterns induced by occupational exposure to irritating chemicals.
Characterisation of Haemophilus influenzae proteins by two‐dimensional gel electrophoresis
Authors: Cash Phillip , Argo Evelyn , Bruce Kenneth D. 
Published: May 2005

AbstractThe proteins of nontypable and type b Haemophilus influenzae isolates were characterised using two‐dimensional polyacrylamide gel electrophoresis (2‐D PAGE). Coomassie Brilliant Blue R‐250 was used for protein detection. Two hundred and twenty eight proteins were resolved from whole cell lysates prepared from a standard nontypable H. influenzae strain (designated HI–64443) when isoelectric focusing was used for the first‐dimensional separation of 2‐D PAGE. When nonequilibrium pH gel electrophoresis (NEPHGE) was used to separate basic proteins in the first dimension, 50 proteins were detected for HI–64443; 20 of the basic proteins detected were considered to be unique for this separation protocol. The apparent molecular weights and isoelectric points were determined for 82 of the proteins resolved for HI–64443. The variation of the proteins from the standard bacterial strain (HI–64443) was determined for nontypable H. influenzae isolates. On the basis of their electrophoretic mobilities, 17.5% of the proteins of HI–64443 were shared by four other non‐typable H. influenzae strains analysed. These data identified both conserved and variable proteins among the nontypable H. influenzae isolates analysed. The results obtained indicated that 2‐D PAGE was able to discriminate nontypable H. influenzae into population clones identified by other procedures. The 2‐D protein profiles obtained for type b H. influenzae strains were similar to those obtained for nontypable H. influenzae strains. The extent of the protein variation observed between type b and nontypable H. influenzae strain was similar to that observed among nontypable strains alone. These data are discussed in relation to the application of 2‐D PAGE as a tool for studies on bacterial epidemiology and for the analysis of the genome structure and gene expression of Haemophilus influenzae.
Single strand conformational polymorphism using capillary electrophoresis with two‐dye laser‐induced fluorescence detection
Authors: Hebenbrock Kirstin , Williams P. Mickey , Karger Barry L. 
Published: May 2005

AbstractCapillary electrophoresis (CE), using a replaceable polymer matrix and two‐dye laser‐induced fluorescence has been applied to single strand conformational polymorphism (SSCP). Two‐dye laser‐induced fluorescence has been used for improved strand identification over a single‐dye approach. Conditions suitable in the capillary format for rapid separation and high resolution have been explored. The influence of separation parameters such as temperature and matrix composition on separation in SSCP was first determined. Short analysis times allowed for fast screening of optimal separation conditions of the sample. Based on these results, the two strands of a standard 255 bp fragment of the lacI gene were resolved within 25 min with replaceable linear polyacrylamide as a separation matrix. The method was then applied to the detection of different mutations, in the presence of wild type, of a 276 bp fragment of the insulin‐like growth factor 1‐binding protein 1 (IGF1‐BP3) gene.
Probing the inner surface of a capillary with the atomic force microscope
Authors: Barberi Riccardo , Giocondo Michele , Bartolino Roberto , Righetti Pier Giorgio 
Published: May 2005

AbstractThe inner surface of a fused silica capillary for zone electrophoresis was probed by the atomic force microscope (AFM) in the contact mode. Only uncoated surfaces were analyzed, after a simple washing cycle with detergent, NaOH and HCl, and final equilibration in distilled water. Three progressively larger surface areas were probed: 0.5 × 0.5 μ, 2 × 2 μm and 4 × 4 μm. In all cases, it was found that the surface is remarkably smooth, with a median height increasing from 1.3 to 5.6 nm; mean height, from 1.3 to 5.8 nm; root‐mean‐squared roughness, from 0.35 to 1.5 nm; and average roughness, from 0.28 to 0.67 nm (the lower values referring to the smaller area, the larger to the largest area probed). The lowest “peaks” detected are of the order of 1–2 nm; some scattered peaks as high as 16 nm are occasionally found. It is concluded that the inner surface of a capillary is not a serrate or notchy structure, but is indeed quite smooth. Since the average roughness is comprised within the thickness of the diffuse double layer (> 10 nm) existing on the silica wall as a result of silanol ionization (and, in fact, it is on the average considerably smaller), it is concluded that it cannot possibly influence peak shape and contribute to peak decay in an electrophoretic run.
Liposome capillary electrophoresis for analysis of interactions between lipid bilayers and solutes
Authors: Zhang Yanxiao , Zhang Rong , Hjertén Stellan , Lundahl Per 
Published: May 2005

AbstractLiposomes, which mimic biomembranes, were used as a pseudostationary phase in capillary zone electrophoresis. The decrease in the mobility of an analyte owing to the presence of liposomes reflected interaction between the analyte and the liposomes. Equations were derived to calculate the specific capacity factor Ks (the capacity factor, K′, normalized to the liposome concentration 1 M) from the migration times and to estimate the difference in free energy, Δ(ΔG0), of the weak analyte/liposome interactions. The order of Ks values for the drugs tested was aspirin < salicylic acid < warfarin < sulfasalazine. The peptide TyrGlySerThrProGlyCysCys interacted more strongly with the liposomes (Ks = 10.1 M−1) than did TyrGlySerThrProGlySerSer (Ks = 9.1 M−1). These results were similar to those obtained earlier by immobilized liposome chromatography.
DNA fingerprinting by hybridisation to oligonucleotide arrays
Authors: Southern E. M. 
Published: May 2005

AbstractMost methods for analysing variation in DNA sequences rely on gel electrophoresis, a method which is difficult to automate and to quantify. Hybridisation to oligonucleotides bound to a solid surface offers alternatives, with the advantage that they can be more readily automated and lend themselves to powerful statistical analytical procedures. In this paper, we discuss a number of different ways in which oligonucleotide arrays could be used for DNA typing for applications in forensic science.
Polymerase chain reaction fingerprinting in fungi using single primers specific to minisatellites and simple repetitive DNA sequences: Strain variation in Cryptococcus neoformans
Authors: Meyer Wieland , Mitchell Thomas G. 
Published: May 2005

AbstractMinisatellites and simple repetitive DNA sequence motifs are used as conventional oligonucleotide probes in DNA‐hybridization‐based fingerprinting. The same oligonucleotides can be used as single primers in the polymerase chain reaction (PCR) to generate individual PCR fingerprints. In this study, the simple repetitive sequences, (CA)8, (CT)8, (CAC)5, (GTG)5, (GACA)4 and (GATA)4, and a minisatellite core sequence derived from the wild‐type phage M13 (5′ GAGGGTGGCGGTTCT 3′) were used as specific, single primers to amplify hypervariable repetitive DNA sequences during PCR analysis. The potential applications of this technique are demonstrated with clinical isolates of the human pathogenic yeest, Cryptococcus neoformans. PCR fingerprint patterns have remained stable after long‐term in vitro passage (>2½ years to date). Hybridization of the primers to blots of electrophoretically separated chromosomes demonstrated that the target sequences recognized by most of the primers are dispersed through the entire yeast genome. Sequence analysis of the cloned bands obtained by PCR fingerprinting indicated that if the same or extremely similar, inversely oriented tandem repeats are located close to each other, when only one repeat‐specific primer is used in the PCR, the region between these repeats is amplified. PCR fingerprinting has a wide range of current and potential applications to fungi, such as clarifying taxonomic questions, facilitating epidemiological studies and improving the diagnosis of mycotic diseases.
Simple repetitive sequences in the genome: Structure and functional significance
Authors: Brahmachari Samir K. , Meera Gopinath , Sarkar Partha S. , Balagurumoorthy Pichumani , Tripathi Jalaj , Raghavan Sowmya , Shaligram Umesh , Pataskar Shashank 
Published: May 2005

AbstractThe current explosion of DNA sequence information has generated increasing evidence for the claim that noncoding repetitive DNA sequences present within and around different genes could play an important role in genetic control processes, although the precise role and mechanism by which these sequences function are poorly understood. Several of the simple repetitive sequences which occur in a large number of loci throughout the human and other eukaryotic genomes satisfy the sequence criteria for forming non‐B DNA structures in vitro. We have summarized some of the features of three different types of simple repeats that highlight the importance of repetitive DNA in the control of gene expression and chromatin organization. (i) (TG/CA)n repeats are widespread and conserved in many loci. These sequences are associated with nucleosomes of varying linker length and may play a role in chromatin organization. These Z‐potential sequences can help absorb superhelical stress during transcription and aid in recombination. (ii) Human telomeric repeat (TTAGGG)n adopts a novel quadruplex structure and exhibits unusual chromatin organization. This unusual structural motif could explain chromosome pairing and stability. (iii) Intragenic amplification of (CTG)n/(CAG)n trinucleotide repeat, which is now known to be associated with several genetic disorders, could down‐regulate gene expression in vivo. The overall implications of these findings vis‐à‐vis repetitive sequences in the genome are summarized.
Novel separation scheme for capillary electrophoresis of enantiomers
Authors: Guttman András 
Published: May 2005

AbstractA systematic approach is described for methods development of chiral separations of weak acidic and basic compounds by capillary electrophoresis, using several natural and derivatized neutral cyclodextrins as chiral selectors. Following the methods development scheme suggested here, the appropriate pH of the running buffer as well as the type and concentration of the cyclodextrin is established for the separation of enantiomers. Preselected chiral selectors of β‐cyclodextrin, γ‐cyclodextrin, hydroxypropyl‐β‐cyclodextrin and dimethyl‐β‐cyclodextrin in low and high concentrations, dissolved in low pH, high pH or pH=pK buffers, are employed during the separation method development and optimization. Depending on the type of separation, introduced by Vigh (desionoselective: only the nondissociated; ionoselective: only the dissociated; duoselective: both enantiomers complex selectively), in most instances at least one of the pH/cyclodextrin combinations results in acceptable separation of the solute enantiomers. The viability of the approach is demonstrated through step by step development of chiral separation for several basic and acidic enantiomers.
Interlaboratory reproducibility of yeast protein patterns analyzed by immobilized pH gradient two‐dimensional gel electrophoresis
Authors: Blomberg Anders , Blomberg Lena , Norbeck Joakim , Fey Stephen J. , Larsen Peter Mose , Larsen Martin , Roepstorff Peter , Degand Hervé , Boutry Marc , Posch Anton , Görg Angelika 
Published: May 2005

AbstractAn interlaboratory comparison was conducted on the positional and quantitative reproducibility of yeast proteins resolved by two‐dimensional polyacrylamide gel electrophoresis (2‐D PAGE) using isoelectric focusing with immobilized pH gradient (pH 4–7) in the first dimension. The basic experimental set‐up was as follows: one laboratory prepared and distributed a [35S]methioninelabeled total yeast protein extract (Göteborg, Sweden), another laboratory prepared the IPG strips to be used by all labs in this study (Munich, Germany), the third laboratory (Aarhus, Denmark) circulated the protocols and coordinated the modest attempts to unify them. Samples were run horizontally in the first dimension and vertically in the second. The gels were sent to Göteborg for processing by phosphoimager technology and computerized image analysis (PDQuest), and the 2‐D PAGE resolved proteins were located and quantified automatically. A subset of 470 spots was manually matched in all gels out of an average of 1328 resolved proteins. The positional interlaboratory comparison revealed great pattern reproducibility, the correlation coefficient in no case being less than 0.9994. In absolute terms an average deviation of 2.8 mm (x‐position) and 1.8 mm (y‐position) were obtained for all nine gels (three gels per lab). The interlaboratory comparison of protein quantitation displayed higher variability, and the best correlation coefficient generated was 0.975. An average standard deviation of 34.5% was calculated for protein quantitation including all three labs, a value slightly higher than the intralaboratory variation (range 20–28%) Thus, despite differences in protocols, chemicals and equipment, the immobilized pH gradient technology gave extremely high positional and quantitative reproducibility. This will greatly facilitate the exchange of data and the establishment of multi‐user image‐based 2‐D gel databases.
Viscous effects in capillary electrophoresis: Theory and experiment
Authors: Schure Mark R. , Murphy Robert E. 
Published: May 2005

AbstractSimple theory which relates the viscous effects of additives used in capillary electrophoresis (CE) to resolution and resolution per unit time are developed for small molecule separations in the absence of sieving effects. The resolution theory shows that there is no advantage to using viscous additives for resolution improvement unless either a binding energy difference exists between the solutes and additive, or viscous effects cause a reduction in the vector sum of electrophoretic and electroosmotic velocities when these are of opposite sign. In general, increasing the viscosity is shown to result in a loss of resolution per unit time. Theory demonstrates that in cases where only binding is considered to be present, the maximum resolution increase is found to be at relatively small amounts of additive. In addition, specific regions of the electrophoreogram may demonstrate resolution enhancement at a specific concentration while other regions exhibit a resolution decrease, as compared to no additive present. CE separations of the components in a Triton surfactant, conducted using a polyethylene glycol (PEG) additive, demonstrate that improved resolution for some of the peaks exists at a specific PEG concentration. These results are mostly explained through the reduction in electroosmotic flow velocity that takes place through additive adsorption to the capillary wall, although some binding is present.
Band width measurement in automated gel electrophoresis apparatus: DNA dispersion in a discontinuous system and in a single buffer
Authors: Yarmola Elena , Chrambach Andreas 
Published: May 2005

AbstractRecent commercial introduction of automated gel electrophoresis apparatus allows for band width measurements during electrophoresis and therefore promises to open up the exploitation of band width and shape for the physical characterization of charged macromolecules in the same manner in which to date quantitative gel electrophoresis had exploited electrophoretic mobility at multiple gel concentrations. The measurements demonstrate decreased band width and therefore increased resolving power for a discontinuous buffer system compared to Tris‐borate EDTA buffer. The dispersion coefficients (D′ = (σ2 ‐σ20)/t) of homogeneous DNA components appear to decrease with gel concentration when either the field strength or the DNA length is small, and increase with gel concentration when these are large. This contrasting response of D′ to increasing gel concentration is presumably due to DNA stretching, which increases in proportion to DNA length and field strength, and to the progressive orientation of agarose with increasing field strength.
Determination of amino acids by on‐line capillary electrophoresis‐electrospray ionization mass spectrometry
Authors: Lu Wenzhe , Yang George , Cole Richard B. 
Published: May 2005

AbstractOn‐line capillary electrophoresis‐electrospray ionization mass spectrometry (CE‐ESMS) has been used for the separation and detection of amino acid mixtures. Four natural amino acids, histidine, tryptophan, phenylalanine, aspartic acid, and a tripeptide, glutathione, were separated and determined. Protonated molecules were detected in the CE‐ESMS mode with detection limits of about one pmol. Optimum CE‐ESMS operating conditions for amino acid analysis were determined employing acetic acid solutions as CE electrolytes. The examined parameters included capillary diameter (50–100 μm internal diameter), applied separation voltage (20–30 kV), and concentration of electrolyte (10–60% acetic acid). Stable working conditions were maintained when the CE currents were less than 18 μA. The use of electrolyte solutions such as those described here instead of true buffer solutions may have advantages for CE‐ESMS systems which employ a “sheathless” interface.