bluuebunny/crossref_metadata_2025_split huggingface dataset:
Unique identification of proteins from small genome organisms: Theoretical feasibility of high throughput proteome analysis
Authors: Cavalcoli James D. , VanBogelen Ruth A. , Andrews Philip C. , Moldover Brian 
Published: May 2005

AbstractWe evaluate current levels of accuracy for estimation of molecular weight (Mr) and isoelectric point (pI) to proteins on two‐dimensional (2‐D) gels as well as the distribution and clustering of proteins in the predicted proteome of E. coli. We also examine the ability to find single candidates within the predicted proteome for matching to a protein seen on 2‐D gels, based on the current level of accuracy. We discuss the levels of accuracy needed to match predicted proteins to observed proteins based solely on Mr and pI criteria obtained from genomic information, and propose methodology to achieve this level of accuracy. In addition, we will address the future goals of this work since the small genomes of bacteria provide a foundation and stepping stone to similar studies in higher organisms.
Melanie II – a third‐generation software package for analysis of two‐dimensional electrophoresis images: I. Features and user interface
Authors: Appel Ron D. , Palagi Patricia M. , Walther Daniel , Vargas J. Reynaldo , Sanchez Jean‐Charles , Ravier Florence , Pasquali Christian , Hochstrasser Denis F. 
Published: May 2005

AbstractAlthough two‐dimensional electrophoresis (2‐DE) computer analysis software packages have existed ever since 2‐DE technology was developed, it is only now that the hardware and software technology allows large‐scale studies to be performed on low‐cost personal computers or workstations, and that setting up a 2‐DE computer analysis system in a small laboratory is no longer considered a luxury. After a first attempt in the seventies and early eighties to develop 2‐DE analysis software systems on hardware that had poor or even no graphical capabilities, followed in the late eighties by a wave of innovative software developments that were possible thanks to new graphical interface standards such as XWindows, a third generation of 2‐DE analysis software packages has now come to maturity. It can be run on a variety of low‐cost, general‐purpose personal computers, thus making the purchase of a 2‐DE analysis system easily attainable for even the smallest laboratory that is involved in proteome research. Melanie II 2‐D PAGE, developed at the University Hospital of Geneva, is such a third‐generation software system for 2‐DE analysis. Based on unique image processing algorithms, this user‐friendly object‐oriented software package runs on multiple platforms, including Unix, MS‐Windows 95 and NT, and Power Macintosh. It provides efficient spot detection and quantitation, state‐of‐the‐art image comparison, statistical data analysis facilities, and is Internet‐ready. Linked to proteome databases such as those available on the World Wide Web, it represents a valuable tool for the “Virtual Lab” of the postgenome area.
Latex allergen database
Authors: Posch Anton , Chen Zhiping , Dunn Michael J. , Wheeler Colin H. , Petersen Arnd , Leubner‐Metzger Gerhard , Baur Xaver 
Published: May 2005

AbstractTwo‐dimensional (2‐D) electrophoresis followed by immunoblotting and N‐terminal protein microsequencing were used to characterize and identify the IgE‐reactive proteins of Hevea latex that are the main cause of the latex type I allergy affecting especially health care workers and spina bifida children. This approach generated a comprehensive latex allergen database, which facilitated the integration of most of the latex allergen data presented in the literature. The major latex allergens Hev b 1, Hev b 3, Hev b 6 and Hev b 7 have been localized on our 2‐D maps. Moreover, we were able to identify six previously undescribed IgE‐binding latex proteins, namely enolase, superoxide dismutase, proteasome subunit C5, malate dehydrogenase, triosephosphate isomerase and endochitinase. The generated latex 2‐D maps will provide valuable information to develop strategies for the isolation of the novel IgE binding proteins in order to study the frequency of sensitization among both risk groups. Detailed knowledge of all proteins involved in latex allergy will allow better diagnosis of latex allergy and to monitor the success of prevention strategies that are needed to reduce the high prevalence of latex allergy among both risk groups.
A two‐dimensional protein map of human amniotic fluid at 17 weeks' gestation
Authors: Liberatori Sabrina , Bini Luca , de Felice Claudio , Magi Barbara , Marzocchi Barbara , Raggiaschi Roberto , Frutiger SÉVerine , Sanchez Jean‐Charles , Wilkins Marc R. , Hughes Graham , Hochstrasser Denis F. , Bracci Rodolfo , Pallini Vitaliano 
Published: May 2005

AbstractUsing updated technical procedures (immobilized pH gradients for isoelectric focusing followed by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis: IPG/SDS‐PAGE) we provide a two‐dimensional (2‐D) map of amniotic fluid (AF) proteins. This map comprises over 800 silver‐stained spots. Over 150 spots have been identified by matching on the net with human plasma and cerebrospinal fluid maps available from SWISS 2DPAGE database; several additional spots were assigned by immunoblotting and/or microanalytical techniques. This report details our investigation on AF proteins focusing on the 17th week of gestation, when AF is most commonly used for clinical evaluation of fetal disorders. As a whole, the map displays a number of potential markers for fetal development and for gestation abnormalities. The 2‐D electrophoretic technique allows the monitoring of all these proteins at the same time along with additional spots that may prove of diagnostic significance.
A novel standardization method for two‐dimensional DNA fingerprints
Authors: Hampe Jochen , Mrowka Ralf , Marczinek Karola , Nürnberg Peter 
Published: May 2005

AbstractTwo‐dimensional (2‐D) DNA fingerprinting is a technique that allows for parallel genome analysis through the simultaneous detection of up to 500 minior microsatellite loci on a 2‐D gel. Separation is performed according to size and melting temperature in the gel. In the application of this technique in genome analysis, a standardized method for the identification of individual spots is required. However, due to the polymorphic nature of up to 80% of the spots, existing standardization methods that have been primarily developed for 2‐D protein patterns are not suitable for this task. We developed a robust method that standardizes 2‐D DNA fingerprint spots on the basis of melting temperature – or denaturing gradient position – and fragment size. An external marker was used as a basis for standardization. A normalization surface was calculated over the gel dimensions by adapting an established numerical iteration technique previously used in physics termed “relaxation method”. The relaxation method works robustly with the irregularly spaced marker spots. The evaluation of the method for a spot of preknown position derived from the TP53 gene revealed a median observed error below 1% for fragment length and denaturing gradient position. The search for candidate minisatellite loci in genomic difference analysis depends on the reliable identification of alleles of this locus in different individuals. We proved experimentally that alleles of a single minisatellite locus cloned from a 2‐D gel cluster on an isothermal line can be reliably identified using the presented standardization method. In conclusion, a standardization tool for a broader application of 2‐D DNA fingerprinting in both tumor analysis and possibly parallel mutation screening is now available.
DNA sequencing with pulsed‐field capillary electrophoresis in poly(ethylene oxide) matrix
Authors: Kim Yongseong , Yeung Edward S. 
Published: May 2005

AbstractDNA sequencing by using pulsed‐field capillary electrophoresis in a linear polymer matrix was performed in this work. The effects of waveform, frequency, modulation depth and heating were investigated. The separation performance for Sanger DNA fragments up to 1000 bp using a pulsed field was compared to isoelectric and isorheic conditions. A separation mechanism in the flexible poly(ethylene oxide) (PEO) gel was also suggested. We found that temperature change generated by either a pulsed field or ultrasound during DNA sequencing can have a deleterious effect on the separation efficiency.
Separation of oligonucleotides of identical size, but different base composition, by free zone capillary electrophoresis in strongly acidic, isoelectric buffers
Authors: Perego Marilena , Gelfi Cecilia , Stoyanov Alexander V. , Righetti Pier Giorgio 
Published: May 2005

AbstractA novel method for analyzing oligonucleotides of the same length, but bearing a single base substitution, is reported, based on free zone capillary electrophoresis (CZE) under rather acidic pH values. For this purpose, a set of four 18‐mers of fairly random base composition has been synthesized, bearing, in nucleotide 9, the following bases: T, C, G or A. Theoretical predictions, based on titration curves of single free nucleotides, allowed us to predict that the simultaneous separation of a mixture of all four oligonucleotides could be possible in a pH 3–4 window. In fact, electrophoresis at pH 5.7 gave a single, asymmetric peak, whereas CZE at pH 4.8 could resolve three out of four species (the T9 and G9 oligonucleotides co‐migrating into a single zone). A unique separation power could be obtained at pH 3.3 in a buffer comprising an amphoteric species (isoelectric iminodiacetic acid, IDA) and 7 M urea. Although IDA exhibited a pI of 2.23 (for a 100 mM solution), the addition of 7 M urea (necessary to denature the oligonucleotides) raised the apparent pH of the solution to 3.3.
Capillary zone electrophoresis of interconverting cis‐trans conformers of peptidyl‐proline dipeptides: Estimation of the kinetic parameters
Authors: Rathore Anurag S. , Horváth Csaba 
Published: May 2005

AbstractPeptides containing proline residues, except at the N‐terminus, exist in cis and trans forms due to the rigidity of the peptidyl‐proline bond. Computer‐simulated and experimental electropherograms have been used to study the interplay of the electrophoretic migration and the kinetics of the cis‐trans interconversion. The measure of the interference by the reaction is the dimensionless Damköhler number (Da) which is the ratio of the migrant's residence time to the characteristic time of reaction in the capillary. The fraction of the trans conformer that is not separable with 90% purity is employed to quantify the extent of overlap between the concentration profiles of the two interconverting migrants and the effect of the various operational variables was examined by simulation. The deterioration of separation, as measured by the degree of overlap, is a quasi sigmoidal function of Da with the selectivity and the intrinsic efficiency of the system as the parameters. At sufficiently low temperatures the interference by reaction kinetics may vanish and the two conformers are separated. At high enough temperatures, when the rate of interconversion is very fast, the two conformers are not separated at all. A simple and rapid method is proposed for estimation of kinetic parameters for the cis‐trans isomerization on the basis of data obtained with phenylalanyl‐proline dipeptide. The procedure involves determination of the equilibrium constant by measuring equilibrium concentrations of the two conformers using nuclear magnetic resonance (NMR) or capillary zone electrophoresis (CZE) at very low temperature, correlation of the peak shapes in the electropherograms with the Da, and finally, evaluation of the forward rate constants from the assigned Da values in the domain: 0.01 < Da < 0.5. Separations using CZE were performed in the temperature range of 1–40°C by using the Beckman P/ACE unit equipped with an auxiliary cooling system. The kinetic data thus obtained showed good agreement (average error less than 5%) with those measured by NMR.
Resolution of the diastereomers of a large synthetic peptide by capillary electrophoresis using nonionic surfactants
Authors: Gilges Martin , Hadley Mark 
Published: May 2005

AbstractA capillary electrophoretic method was developed for the determination of the diastereomeric composition of the bradykinin B2 antagonist, SB‐238592‐DB. The nonionic surfactant Brij 35 was demonstrated to be suitable for the baseline resolution of the three diastereoisomers of SB‐238592‐DB in the presence of a low pH phosphate buffer and small volume percentages of acetonitrile.
Identification of plasma proteins facilitated by enrichment on particulate surfaces: Analysis by two‐dimensional electrophoresis and N‐terminal microsequencing
Authors: Lück Martin , Schröder Werner , Harnisch Stephan , Thode Kai , Blunk Torsten , Paulke Bernd‐R. , Kresse Mayk , Müller Rainer H. 
Published: May 2005

AbstractPlasma protein adsorption on intravenously injectable drug carriers is regarded as an important factor for the fate of the particles in the body after their administration. Therefore, the plasma protein adsorption patterns on a number of different carrier systems were analyzed in vitro employing two‐dimensional electrophoresis (2‐DE). The particulate systems presented in this study were polystyrene (PS) model particles, PS nanoparticles surface‐modified by adsorption of a surfactant, a commercial fat emulsion, and magnetic iron oxide particles used as contrast agents in magnetic resonance imaging. Most of the spots in the plasma protein adsorption patterns could be identified by matching the resulting 2‐DE gels with a reference map of human plasma proteins. Several other proteins that indicated preferentially adsorbed proteins on the surface of the particles investigated have either not been identified on the reference map, or their identity was found to be ambiguous. The relevant proteins are all present in plasma in low abundance. Since these proteins were strongly enriched on the surface of the particles, the resulting spots on the 2‐DE gels were successfully identified by N‐terminal microsequencing. With this approach, two chains of spots, designated PLS:6 and PLS:8, were determined on a plasma reference map: inter‐α‐trypsin inhibitor family heavy chain‐related protein (also named PK‐120) and a dimer of fibrinogen γ, respectively. Plasma gelsolin is presented in a 2‐DE adsorption pattern of PS model particles. One of the main proteins adsorbed by droplets of a commercial fat emulsion was identified as apoliprotein H. Moreover, the positions of apolipoproteins apoC‐II and apoC‐III were also verified on the 2‐DE protein map of human plasma. Thus, protein adsorption experiments of the kind presented in this study are increasing our insight into human plasma proteins.
Capillary ion analysis of potassium concentrations in human vitreous humor
Authors: Ferslew Kenneth E. , Hagardorn Andrea N. , Harrison M. Travis , McCormick William F. 
Published: May 2005

AbstractCapillary ion analysis (CIA) is a form of capillary electrophoresis which uses the differential electrophoretic mobility of ions to perform a separation of an ionic mixture. Application of this technique for direct detection of potassium concentrations in human vitreous humor was the purpose of this investigation. CIA was performed using a Waters Quanta 4000 Capillary Electrophoresis System with a 745 Data Module using a 75 μm × 60 cm capillary and a run electrolyte of 67.7 mg hydroxyisobutyric acid (HIBA), 52.8 mg 18‐crown‐6 ether and 64 μL UV‐CAT‐1 reagent (4‐ethylbenzylamine) in a volume of 100 mL water (18 Mohm) with a voltage of 20 kV using ultraviolet absorption detection at 214 nm. Migration times were: ammonium ion, 2.86 min; potassium, 3.24 min; calcium, 3.84 min; sodium, 3.98 min; barium (internal standard), 4.68 min; and lithium, 4.79 min. Correlation coefficients (r) between peak area ratios and concentration ranges of 2.5–144 mmole/L (100–1000 ppm) were from 0.9855 to 0.9999. Coefficients of variation (CV) ranged from 1.45 to 13.8% between days and from 1.38 to 9.43% within‐day. Application of this methodology to twenty‐five vitreous humor specimens from forensic cases was compared to analysis by ion‐specific electrode for potassium concentration. Comparison of CIA to ion‐specific electrode analysis of vitreous humor potassium concentrations revealed a correlation coefficient of 0.9642. CIA is applicable to forensic analysis of potassium concentration in forensic vitreous humor specimens. Quantitation of numerous cation concentrations is possible by direct CIA of vitreous humor.
Strategies for the capillary electrophoretic separation of indole alkaloids in Psilocybe semilanceata
Authors: Pedersen‐Bjergaard Stig , Rasmussen Knut Einar , Sannes Ellen 
Published: May 2005

AbstractWhile the hallucinogenic mushrooms Psilocybe semilanceata have previously been analyzed for the indole alkaloids psilocybin and baeocystin by capillary zone electrophoresis (CZE) at pH 11.5, the present work focused on the development of an alternative and complementary capillary electrophoretic method for their identification. Owing to their structural similarity and zwitterionic nature, the compounds were difficult to resolve based on different interactions with cationic or anionic micelles. However, while the attempts with micellar electrokinetic chromatography (MEKC) were unsuccessful, rapid derivatization with propyl chloroformate and reanalysis by CZE at pH 11.5 was effective to support identification of the two indole alkaloids. Psilocin was difficult to analyze by CZE at pH 11.5 owing to comigration with the electroosmotic flow. For this compound, the pH of the running buffer was reduced to 7.2 to effectively enhance the electrophoretic mobility.
The analysis of fountain pen inks by capillary electrophoresis with ultraviolet/visible absorbance and laser‐induced fluorescence detection
Authors: Rohde Ellen , Vogt Carla , Heineman William R. 
Published: May 2005

AbstractInformation on the identity of inks adds to the circumstantial evidence in legal cases involving fraudulent documents. In combination with optical methods, multiple thin‐layer chromatography (TLC) is currently the analytical tool used by forensic chemists to separate, compare and distinguish inks based on their dye composition. In our studies, capillary electrophoresis (CE) was used for the analysis of water‐soluble fountain pen inks. Inks are complex mixtures of synthetic organic and inorganic dyes, surfactants, resins and other components. The investigations included the development of an electrophoretic separation method, the optimization of an extraction procedure for inks from paper as well as the evaluation of ultraviolet/visible (UV/VIS) absorbance and laser‐induced fluorescence (LIF) detection for the analysis of inks by CE. Good results for the separation of 17 blue and black inks of different manufacturers and countries of origin were obtained with 100 mM borate buffer, pH 8.0, containing 20% methanol. The electropherograms of the inks and their extracts from paper showed patterns that were in most cases distinctly different from each other. Ultraviolet/visible scans can be used to compare spectra of the separated main and trace components of inks. Fluorescence detection at different excitation and emission wavelengths was more sensitive, but added to the complexity of the electropherograms due to the excitation of coextracted fluorescing paper components. The resolution power of CE combined with the information content provided by the detection modes investigated prove CE to be a powerful tool for the identification of water‐soluble inks used on paper documents.
DNA fingerprinting of Listeria monocytogenes using enterobacterial repetitive intergenic consensus (ERIC) motifs‐polymerase chain reaction/capillary electrophoresis
Authors: Sciacchitano Carl J. 
Published: May 2005

AbstractThe molecular technique, enterobacterial repetitive intergenic consensus (ERIC)‐polymerase chain reaction (PCR) produces genomic DNA fingerprint that discriminate bacterial species and strains. This technique was applied to the characterization of Listeria monocytogenes, an important food‐borne pathogen implicated in numerous cases of listeriosis. The ERIC‐PCR resulted in distinct DNA fingerprinting patterns of all L. monocytogene serotypes and Listeria species. Analysis of the genomic DNA fingerprints was accomplished using capillary electrophoresis (CE), an alternative technique to the conventional agarose gel method. The optimization of CE conditions (electrokinetic injection, applied voltage) resulted in the resolution of amplified DNA fragments up to 1000 bp. Comparisons of electropherograms provided genomic fingerprint templates which could be further used for supplementary information. The ERIC‐PCR method coupled to CE provides a rapid technique in differentiating bacterial spp., and may contribute relevant information in food‐borne outbreak studies.
Analysis of the D1S80 locus by capillary electrophoresis
Authors: Mitchell Joseph W. , Walsh David 
Published: May 2005

AbstractWe have demonstrated in a systematic manner that the capillary electrophoresis (CE) method of genotyping the human D1S80 locus is an effective replacement for the commonly used gel electrophoresis method. The CE method is fast, with a total run time of less than 22 min per sample. Separation has been optimized so that resolution for all pairs of alleles from 14 through 41 is greater than 1, providing unambiguous identification. The method utilizes run buffer containing 0.30% hydroxyethyl cellulose as the sieving polymer in a 50 μm internal diameter (ID) column coated with a 0.1 μm DB‐17 film. Laserinduced fluorescence (LIF) with YO‐PRO‐1 intercalating dye is used for detec‐tion. Internal standards of 300 and 1000 bp bracket the DlS80 region in the electropherogram. Two typing procedures were evaluated: matching of the normalized migration times of sample alleles to ladder alleles; and software alignment of sample and ladder electropherograms using the internal standard peaks as references. Using the first procedure, 91.5% of the D1S80 genotypes were unambiguous, while 100% were unambiguous using the second procedure. Seventy‐nine routine samples were analyzed in a side‐by‐side comparison of the CE and slab gel methods, with complete agreement of the results. Twenty‐two samples were selected from a large database previously analyzed by the slab gel method to demonstrate that all alleles from 14 through 41 were typed correctly, including samples containing adjacent alleles. Additionally, 43 samples containing alleles greater than the 41 repeat number were typed correctly.
Nonradioactive sequence‐tagged microsatellite site analyses: A method transferable to the tropics
Authors: Lagoda Pierre J. L. , Dambier Dominique , Grapin Agnès , Baurens Franc‐Christophe , Lanaud Claire , Noyer Jean‐Louis 
Published: May 2005

AbstractUtilization of existing isozyme analysis facilities to detect sequence‐tagged microsatellite site (STMS) polymorphism or any simple sequence repeat (SSR) variation is described. Different parameters concerning the difficulties in transferring molecular techniques to less sophisticated laboratory infrastructures (i. e. tropical outstations) are discussed (e.g. reproducibility, efficacy, precision). Nonradioactive STMS analysis is bound to foster collaborative research between “biodiversity” and “biotechnology” centers.