bluuebunny/crossref_metadata_2025_split huggingface dataset:
The effect of blob size and network dynamics on the size‐based separation of polystyrenesulfonates by capillary electrophoresis in the presence of entangled polymer solutions
Authors: Cottet Hervé , Gareil Pierre , Viovy J.‐L. 
Published: May 2005

AbstractThis work focuses on the separation of standard polystyrenesulfonates (PSS), with molecular masses (Mr) between 16 and 990 × 103 in capillaries filled with semidilute (entangled) linear hydrophilic polymers. Contrary to cross‐linked chemical gels, which produce permanent networks, solutions of linear polymers lead to dynamic networks. The analytical performances and migration mechanisms are discussed on the basis of experiments performed in solutions of linear polyethyleneoxides and derivatized celluloses of various molecular masses. The influence of the mesh size and of the lifetime of the obstacles of the separating network has been investigated in detail. The mesh size is assimilated to the blob size of the separating polymer and is a decreasing function of its concentration. The lifetime of the obstacles of the network, identified with the reptation time of the polymer chain, characterizes its dynamics. This characteristic time increases with both the molecular weight of the separating polymer and its concentration. Its impact was first examined at fixed blob size. Then, the influence of the blob size was studied while keeping the reptation time of the network constant. By doing so, the existence of interactions between the solute and the separating polymer or between the solute and capillary wall can be more safely assessed. It appears that the reptation time of the mesh has a large influence on the electrophoretic mobility of the PSSs under a threshold value, which is of the order of magnitude of the time taken by the PSS to migrate on the blob size. Also shown are separations using networks made up with mixtures of polyethyleneoxides of the same nature and same mass concentration, but of very different molecular masses. This latter approach allows one to adapt the viscosity of the solution and the dynamics of the network, keeping the blob size constant.
Analysis of antisense oligonucleotides by on‐capillary isotachophoresis and capillary polymer sieving electrophoresis
Authors: Khan Kamran , Van Schepdael Ann , Hoogmartens Jos , Saison‐Behmoaras Tula , Van Aerschot Arthur 
Published: May 2005

AbstractAn attempt was made to evaluate the stability of an antisense oligonucleotide against nucleases present in HBL 100ras cells. To detect nanomolar concentrations of the oligonucleotide, a sensitive detection system was required. A combination of capillary electrophoresis/laser‐induced fluorescence (CE‐LIF) with fluorescence derivatization did not improve the sensitivity significantly and also resulted in loss of separation of the derivatized sample. On‐column isotachophoresis for the preconcentration of oligonucleotide samples in DB‐17 coated capillaries filled with hydroxyethyl cellulose solution could be an alternative. The isotachophoresis (ITP) step allows injection of up to 40% of the capillary volume without loss in peak resolution and peak efficiency. Using ITP‐capillary polymer sieving electrophoresis (CPSE), the limit of quantitation at a signal‐to‐noise ratio of 10 was 73 ng/mL for a 12‐mer oligonucleotide. Using these conditions, the gain in sensitivity was 125.
Subnanomolar detection limit for sodium dodecyl sulfate — capillary gel electrophoresis using a fluorogenic, noncovalent dye
Authors: Harvey Michael D. , Bandilla Dirk , Banks Peter R. 
Published: May 2005

AbstractPicomolar limits of detection are obtained using the noncovalent, fluorogenic dye, Sypro Red. The size separation of four commonly used sodium dodecyl sulfate ‐ capillary gel electrophoresis (SDS‐CGE) molecular weight markers with 8% linear polyacrylamide (PAA) as the sieving matrix is used to construct a calibration curve for molecular weight determinations. SDS‐CGE purity and molecular weight determination of purified chorismate mutase‐prephenate dehydrogenase (CMPD) from Escherichia coli is shown to be comparable in accuracy with slab gel SDS‐polyacrylamide gel electrophoresis (SDS‐PAGE). A migration time precision study indicates excellent reproducibility. Sypro red labeling of SDS‐bovine serum albumin (SDS‐BSA) complexes at nanomolar protein concentrations suggests assay detection limits surpassing those of silver staining. This detectability exceeds that achieved in previous SDS‐CGE laser‐induced fluorescence studies. This approach is expected to be easily adapted for use with commercial polymer formulations and automated instrumentation.
Sodium dodecyl sulfate‐capillary electrophoresis of proteins in a sieving matrix utilizing two‐spectral channel laser‐induced fluorescence detection
Authors: Craig Douglas B. , Polakowski Robert M. , Arriaga Edgar , Wong Jerome C. Y. , Ahmadzadeh Hossein , Stathakis Costas , Dovichi Norman J. 
Published: May 2005

AbstractWe report a method for protein labeling, separation by capillary electrophoresis in a polymer sieving matrix, and detection by laser‐induced fluorescence. Different dyes are used to label standard and sample proteins. A two‐spectral channel detector resolves fluorescence from the sample and standards. Comparison of the migration time of the sample and standards permits the precise determination of molecular weight, irrespective of variations in run‐to‐run migration times.
Enhanced sensitivity for sequence determination of major histocompatibility complex class I peptides by membrane preconcentration ‐ capillary electrophoresis ‐microspray ‐ tandem mass spectrometry
Authors: Naylor Stephen , Ji Qinchung , Johnson Kenneth L. , Tomlinson Andy J. , Kieper William C. , Jameson Stephen C. 
Published: May 2005

AbstractSequence analysis of antigenic major histocompatibility complex (MHC) class I peptides requires minimizing sample loss and enhancing mass spectrometric sensitivity. In order to facilitate such analyses, we have coupled on‐line membrane preconcentration‐capillary electrophoresis (mPC‐CE) with microspray mass spectrometry (mPC‐CE‐μMS) and tandem mass spectrometry (mPC‐CE‐μMS/MS). Specifically, cell lysate from ∼ 109 EG‐7 mouse tumor cells was immunoprecipitated and the released MHC class I peptides were subjected to reverse‐phase HPLC. An HPLC fraction containing antigenic peptide(s) shown to induce T‐cell stimulation was subjected to mPC‐CE‐μMS. Approximately 10 μL (from 100 μL) of the fraction was pressure‐injected and concentrated on a styrenedivinylbenzene (SDB) impregnated membrane. The peptides were eluted from the membrane with ∼100 nL of 80% methanol, sandwiched between a leading stcking buffer (LSB, also serving as CE separation medium) of ∼110 nL of 0.1% acetic acid in 10% methanol, and a trailing stacking buffer (TSB) of ∼ 110 nL of 0.1% NH4OH. On application of the CE voltage the peptides are subjected to moving boundary transient isotachophoresis and focused. The peptides were separated in a Polybrene‐coated capillary with application of −20 kV in reverse polarity mode and subsequently sprayed via an emitter coupled to the CE capillary by a liquid junction containing a platinum wire. An ion at m/z 482.3 was detected and subjected to mPC‐CE‐μMS/MS and determined to be SIINFEKL, a peptide (OVA) known to be antigenic in the mouse model system. Sensitivity enhancement over conventional mPC‐CE‐MS and MS/MS was ∼100‐fold.
Fluorescence imaging of light absorption for axial‐beam geometry in capillary electrophoresis
Authors: Johansson Jonas , Johansson Thomas , Nilsson Staffan 
Published: May 2005

AbstractA new method for investigation of axial‐beam absorption detection for improved detection limits in microcolumn separations is reported. The method is based on fluorescence imaging of light absorption along a separation capillary. The probing UV light is introduced at one end of the capillary and shows an exponential fall‐off along the capillary. As the UV light propagates through the sample peaks, an additional loss in intensity will be observed. In order to view the absorption profile along the capillary, a background fluorophore is added to the buffer. A charge‐coupled device (CCD) detector and imaging optics are placed beside the capillary to view the capillary in a direction perpendicular to the capillary. Signal integration is employed for consecutive exposures as well as for neighboring detector pixels in order to increase the signal‐to‐noise ratio. Measurements for stilbene 3 with sulforhodamine B as a background fluorophore are presented. The characteristics of the detection method and potential improvements are discussed.
A computer‐controlled variable light attenuator for protection and autoranging of a laser‐induced fluorescence detector for capillary zone electrophoresis
Authors: Merz Jerry M. , Mort Andrew J. 
Published: May 2005

AbstractPhotodetectors have a limited range over which they can measure light intensities for any particular setting. The intensity of light reaching the detector can be kept within this range by using a liquid crystal variable light attenuator controlled by a computer that continuously checks the amount of light reaching the detector and adjusts the attenuation to an appropriate level. Using such a system we have constructed an intensified charge‐coupled device (ICCD) camera‐based detector with a dynamic range of over six orders of magnitude which is never exposed to damaging or saturating light levels.
Dual UV‐absorbing background electrolytes for simultaneous separation and detection of small cations and anions by capillary zone electrophoresis
Authors: Xiong Xiang , Li Sam F. Y. 
Published: May 2005

AbstractThe simultaneous separation and detection of small cations and anions by capillary zone electrophoresis (CZE) with indirect ultraviolet (UV) detection was successfully demonstrated in a background electrolyte (BGE) containing two UV‐absorbing components. Benzylamine, imidazole, benzenesulfonic acid, sulfosalicylic acid, and pyromellitic acid were tested as the components of the BGE. The success of the simultaneous separation of the cations and anions is dependent upon the proper selection of the electrolyte components and control of the migration of the ions towards the detector. High pH is beneficial to the detection of anionic analytes but not to the separation of cationic analytes because of large electroosmotic flow produced under this condition. The upper pH limit of the working pH range is confined by the pKa value of the cationic component of the BGE. The influence of pH and total electrolyte concentration on the electroosmatic flow (EOF) counteracted each other. This counteraction effect imposes an upper limit on the change of total electrolyte concentration at certain pH. It was found that the EOF should be larger by at least 10 × 10−5 cm2V−1V1 than the electrophoretic mobilities of the anions so that the anions could be detected on the cathodic side within reasonable times and with good peak shapes. In the imidazole‐sulfosalicylic acid BGE, the detection limits (signal to noise, S/N = 3) for the cations and anions ranged from 100 to 900 ppb. In the benzylamine‐pyromellitic acid BGE, K+, Na+, Li+, CH3 COO−, HPO42−, F−, C1O3−, C1O4−, NO3−, NO2−, Cl− and SO42− were separated within twelve minutes. The strategies for selection of the electrolyte components of the binary BGE were also discussed.
Capillary electrophoresis/laser‐induced fluorescence detection of fluorescein as a groundwater migration tracer
Authors: Ferguson Patrick L. , Grange Andrew H. , Brumley William C. , Donnelly Joseph R. , Farley John W. 
Published: May 2005

AbstractCapillary electrophoresis (CE) has been applied to the determination of the groundwater migration tracer dye fluorescein based on laser‐induced fluorescence (LIF) detection and compared to determinations obtained with traditional spectrofluorimetry. Detection limits of injected dye in the low parts per trillion (ppt) ranges have been accomplished with both CE/LIF based on the Ar ion laser and with a spectrofluorimeter. This approach was used for a real‐world problem in determining groundwater migration between adjacent Resource Conservation and Recovery Act (RCRA) and Superfund sites by the Environmental Sciences Division in response to regional needs and as application of new analytical tools under development. Fluorescent dye was injected into source wells and then was determined in monitoring wells by extracting pads that adsorbed the dye or by directly determining the dye in the water using solid‐phase extraction (SPE), a preconcentration technique. The approaches based on CE/LIF exhibits increased specificity over existing approaches due to the separation and unique migration time of the dye. Additional studies were aimed at achieving sub‐ppt levels in the water using solid‐phase extraction and field‐amplified injection techniques.
Steady‐state electrolysis of an ampholyte solution and possibility of violation of the “law of pH monotony”
Authors: Stoyanov Alexander V. , Righetti Pier Giorgio 
Published: May 2005

AbstractThe problem of stationary electrolysis of a single‐component ampholyte solution is analyzed. The effect of nonconstant relative mobility is taken into account. It is demonstrated that an incorrect dissociation model (i.e., expressing the resulting ampholyte flux by the arithmetic sum of the fluxes for “cationic” and “anionic” species) may lead to the violation of the so‐called “law of pH monotony”.
Recent developments in capillary isoelectric focusing with whole‐column imaging detection
Authors: Fang Xiaohong , Tragas Charalambos , Mao Qinglu , Pawliszyn Janusz , Wu Jiaqi 
Published: May 2005

AbstractCapillary isoelectric focusing (CIEF) is a high resolution technique for protein separation. The on‐column single point detector requires a mobilization step which lengthens the analysis time and causes an uneven resolution along the separation column. The real time and whole column imaging detection has been developed for performing CIEF without mobilization. Three types of imaging detection systems have been developed: optical absorption, refractive index gradient, and laser induced fluorescence. This technique provides a fast analysis speed (about 6 min) and a good resolution of 0.03 pH unit level. Using the absorption imaging detector, ampholyte‐free IEF in tapered capillary is being demonstrated, which eliminates the interference of the expensive carrier ampholytes for protein detection in UV region. Recent advancements in this imaged CIEF technique as well as its applications are reviewed.
Optimization of solid phase microextraction ‐ capillary zone electrophoresis ‐ mass spectrometry for high sensitivity protein identification
Authors: Figeys Daniel , Zhang Yanni , Aebersold Ruedi 
Published: May 2005

AbstractWe have previously described the use of a solid phase extraction (SPE) ‐ capillary zone electrophoresis (CZE) ‐ tandem mass spectrometry (MS/MS) system for protein analysis at the low femtomole to subfemtomole level. Here we describe the systematic optimization of a number of parameters which facilitate the use of the SPE‐CZE‐MS/MS system and further enhance its performance. Specifically, we describe a robust SPE cartridge design which can be assembled without the use of glue, the evaluation of procedures to chemically modify the inner wall of the fused‐silica capillaries used in the system to improve separation and reproducibility, and the comparison of different reverse‐phase (RP) resins used for the SPE cartridge. We also explored the effects of transient isotachophoresis with respect to system performance and compatibility with different fused‐silica surface coatings, the RP resins used, and MS/MS. The enhanced performance of the optimized system is demonstrated by the analysis of calibrated tryptic digests of bovine serum albumin (BSA).
Analysis of sequence homogenisation in rDNA arrays of Haemonchus contortus by denaturing gradient gel electrophoresis
Authors: Gasser Robin B. , Zhu Xingquan , Chilton Neil B. , Newton Lisa A. , Nedergaard Trine , Guldberg Per 
Published: May 2005

AbstractTesting different theories of concerted evolution experimentally has been hampered mainly due to the lack of appropriate model systems and technical limitations. In this study, we employed a denaturing gradient gel electrophoresis (DGGE) approach for the display and definition of nucleotide variations in the second internal transcribed spacer (ITS‐2) of ribosomal DNA (rDNA) of the parasitic nematode, Haemonchus contortus. The ITS‐2 was amplified from individual adult nematodes by PCR and subjected to DGGE. Of the 94 individuals (representing nine different populations) analysed, 13 different DGGE profiles were displayed. Eighteen bands representing those profiles were excised and sequenced. Sequencing defined 13 different types of ITS‐2 with 12 nucleotide variations (4 transitions, 5 transversions, 1 insertion and 2 deletions) which could be related to particular positions of the predicted secondary structure for the ITS‐2 pre‐rRNA. The results showed that individuals of interbreeding populations of H. contortus can have rDNA arrays that are partially or fully homogenised for different sequence variants (despite interindividual variation), suggesting that the homogenisation process is driven mainly by intrachromosomal exchange. The findings also demonstrated the capacity of the DGGE‐sequencing strategy to quantify the frequency of ITS‐2 sequence types within individual nematodes from different populations without the need for cloning or Southern blot procedures. This has important implications for studying the mechanisms of sequence homogenisation in rDNA and pre‐rRNA processing as well as for elucidating speciation events and population differentation at the molecular level.
Short tandem repeat (STR) locus HUMD8S306 in a large population sample from Germany
Authors: Benecke Mark , Knopf Manuela , Voll Wolfgang , Oesterreich Wolfgang , Jacobi Yvonne , Edelmann Jeanett 
Published: May 2005

AbstractApplied DNA typing in medico‐legal investigations, in criminalistic practice, and in paternity cases often relies on high inclusion and exclusion probabilities. For that reason, the short autosomal tandem repeat locus D8D306 was validated for forensic use and incorporated into a nonoverlapping multiplex reaction with HUMDHFRP2 and HUMCD4. The allele frequencies of D8S306 in four different regions of Germany (n = 1220 alleles) were determined for use in a population database; the allele distributions did not significantly deviate from each other. The hererozygosity of D8S306 is 83%, expected exclusion chance in stain cases is 96% (paternity cases: 69%), the lowest amount of sucessfully amplified DNA was 30 pg. The alleles are in Hardy‐Weinberg equilibrium (exact test).
Rapid fluorescent monitoring of total protein patterns on sodium dodecyl sulfate‐polyacrylamide gels and Western blots before immunodetection and sequencing
Authors: Alba F. Javier , Daban Joan‐Ramon 
Published: May 2005

AbstractThe fluorogenic dye 2‐methoxy‐2,4‐diphenyl‐3(2H)‐furanone (MDPF) has been used for the detection of total protein patterns on polyvinylidene difluoride (PVDF) membranes. Fluorescent staining of protein bands on membranes with this covalent dye is completed in 20 min. Wet membranes are translucent, allowing protein visualization by transillumination with ultraviolet light. The resulting images can be recorded using Polaroid film or a charge‐coupled device camera. Electrophoretic bands containing 5–10 ng of protein can be detected on the MDPF‐stained Western blot. When proteins are directly transferred to the membrane using a slot blotting device, as little as 0.5 ng of protein can be detected. Previous visualization of protein bands on sodium dodecyl sulfate‐polyacrylamide gels with the noncovalent fluorescent dye Nile red (Alba et al., BioTechniques, 1996, 21, 625–626) does not interfere with further MDPF staining and fluorescent detection of these bands transferred to PVDF membranes. Thus, Nile red and MDPF staining can be performed sequentially, allowing the rapid monitoring of total protein patterns on both the electrophoretic gel and Western blot. Using the conditions described in this study, MDPF staining does not preclude further N‐terminal microsequencing and immunodetection of specific bands with polyclonal antibodies.
An electrophoretic study on interactions of albumins of different species with immobilized Cibacron Blue F3G A
Authors: Miller Ingrid , Gemeiner Manfred 
Published: May 2005

AbstractAlbumins of different species, varying in electrophoretic mobility, were compared in their interaction with the dye Cibacron Blue F3G A. Immobilized by coupling to a high molecular weight dextran (“blue dextran”), the dye was used as a ligand in affinity electrophoresis in different setups. One‐dimensional electrophoresis with blue dextran entrapped in an intermediate gel and two‐dimensional applications with transverse gradients (affinity titration curves, zonal electrophoresis in linear ligand gradients) were performed. Compared to the human homologue, animal albumins display more complex patterns and interaction profiles, depending on pH and ionic strength of the buffers. Results may differ considerably from those obtained by affinity chromatography, illustrating the additional screening potential of the electrophoretic methods. Comparison of different samples under the influence of ligand competition, reducing conditions, or denaturing agents gives supplementary information on conformational behavior of the proteins.
DNA preparation and efficient microsatellite analysis from insect hemolymph
Authors: Gerken Thomas , Kurtz Joachim , Sauer Klaus P. , Lubjuhn Thomas 
Published: May 2005

AbstractA simple and time‐saving method for DNA preparation for efficient microsatellite analysis is described. The method is based on thermal treatment of only 1–5 μL of insect hemolymph in a Chelex 100‐suspension. Since hemolymph withdrawal does not harm the insects, analysis of mating systems, population structure and phylogenetic reconstruction can be conducted with minimal experimental influence.
Identification of animal species in ground meat mixtures by multivariate analysis of isoelectric focusing protein profiles
Authors: Skarpeid Hans‐Jacob , Kvaal Knut , Hildrum Kjell Ivar 
Published: May 2005

AbstractThe present work investigates the possibility of constructing a multivariate calibration model to predict the composition of ground meat with respect to animal species, based on intensity profiles from isoelectric focusing of water‐soluble proteins in mixtures of ground meat. Samples containing various amounts of beef, pork and turkey meat were analyzed by isoelectric focusing in immobilized pH‐gradients. The gels were photographed and the images transferred to a digital format. The digital images were processed in order to reduce background and optimize signal strength. Resulting gel profiles were analyzed by multivariate regression allowing the determination of sample composition with prediction errors close to 10%.