bluuebunny/crossref_metadata_2025_split huggingface dataset:
Electrophoresis in gels containing zwitterionic groups
Authors: Hjelmeland Leonard M. , Allenmark Stig , An der Lan Birgit , Jackiw B. Andrew , Nguyen Nga Y. , Chrambach Andreaas 
Published: May 2005

AbstractA zwitterionic methacrylamide derivative, (3‐sulfopropyl)dimethy1(3‐methacrylamidopropyl) ammonium inner salt (MAPS), was synthesized and then copolymerized with N,N′‐methylenebisacrylamide (Bis) to make gels carrying covalently bound sulfobetaine groups. Gels were also prepared with the addition of (3‐sulfopropyl)trimethyl ammonium inner salt (TMAPS) to Bis‐crosslinked polyacrylamide prior to polymerization. The properties of these gels in gel electrophoresis and electrofocusing were tested and compared with those of Bis‐crosslinked polyacrylamide.In MAPS gels the absolute migration velocities of proteins and dyes at either polarity of migration were decreased. Anionic species were much more strongly retarded than cationic species. Ferguson plots (based on protein mobilities relative to dye in a continuous buffer) of catalase at pH 4 obtained on MAPS gels had a similar slope but lower y‐intercept compared with those on equivalent polyacrylamide gels, indicating that the available (effective) net charge on the protein was decreased because it interacts with the zwitterionic charge on the gel. Similarly, the addition of TMAPS to polyacrylamide decreased the relative catalase mobility at non‐restrictive gel concentrations. However, at gel concentrations above 4 %T, relative mobilities were increased, presumably because the effective pore size of the gel was increased through inhibition of polymerization by the zwitterionic compound.MAPS gels did not exhibit electroendosmosis by the criterion of cyanocobalamin displacement. Resolution between peptides in MAPS gels, and between proteins in ternary copolymer gels made of MAPS, Bis and polyacrylamide, was similar to that on polyacrylamide. Bands of catalase appeared sharpened in MAPS gels. pH Gradients form, decay and exhibit conductance gaps in MAPS electrofocusing gels and in polyacrylamide gels in presence of 0.25‐1.5 M TMAPS, in a manner which was qualitatively indistinguishable from polyacrylamide.
Electrophoresis of viral proteins in formic acid 1. General aspects
Authors: Heukeshoven Jochen , Dernick Rudolf 
Published: May 2005

AbstractThe excellent solublizing properties of high concentrations of formic acid solutions for polypeptides and proteins, including those with high hydrophobicity, are used in polyacrylamide gel electrophoresis. The technical handling of the system is described in detail. The structural polypeptide of poliovirus type1, VP1, VP2, VP3 an dVP4, until now separable in polyacrylamide gel rods containing 70% formic acid. By two‐dimensional analysis, namely formic acid electrophoresis in the first dimension and SDS electrophoresis in the second, we have shown that the electrophoretic mobility in formic acid is of the order VP2‐VP1‐VP3‐VP4. with several marker proteins we have also demonstrated that the migration velocity depends not only on the molecular weights but also on the secondary structure of the proteins, especially on disulfide cross‐links. High concentrations of salts (3 MCsCI or 6 m Guanidine hydrochloride) in the protein sample do not change the separation pattern, which, however, is strongly affected by 6 m urea.
Peptide maps of labeled poliovirus proteins after two‐dimensional analysis by limited proteolysis and electrophoresis in sodium dodecyl sulfate
Authors: Wiegers Klaus J. , Dernick Rudolf 
Published: May 2005

AbstractA simple technique for characterization and identification of proteins after high resolution two‐dimensional (2D) electrophoresis is described. Poliovirus proteins were separated by 2D analysis and then subjected to llimited proteolysis in sodium dodecyl sulfate (SDS) by alfa‐Chymotrypsin or Staphylococcus V8 protease. The cleavage products were analyzed by SDS‐polyacrylamide gel electrophoresis (SDS‐PAGE). The pattern of peptides obtained was characteristic for a given protein and for the protease used and was highly reproducible. Alfa‐Chymotrypsin produced fewer and larger peptides and was suitable for large poliovirus proteins. Staphylococcus V8 protease yielded smaller peptides and therefore showed a higher degree of discrimination between the poliovirus proteins analyzed. A precursor product relationship for a number of poliovirus proteins was determined by this simple technique.
Anamalous electrophoresis behavior of rebonuclease from human urine and a novel treatment method for reliable analysis
Authors: Thomas John M. , Hodes Marion E. 
Published: May 2005

AbstractSince Desialylated rebonuclease (RNase) from human urine has a highle alkaline isoelectric point (pI 9.5–10.5), an acidic buffer system is required for electrophoretic analysis of different urine specimens. In a common acid disc electrophoresis system in thin‐layer polyacrylamide gel, the cathodal mobility of the RNase enzyme(s) was markedly reduced from that expected on the basis of the estimated pI. Dialysis urine against water had a very detrimental effect on the separation of the isozymes, and the addition of buffer to the dialyzed urine further exacerbated this effect. The urin RNase exhibited anomalous electrophoretic behavior in the form different urine specimens or from the same urine specimen adjusted to different concentrations. Treatment of urine with various cations had a significant, positive effect on the migration and resolution of the isozymes. This and other experimental evidence is consistent with the hypothesis that the anomalous behavior was due to the binding of anions from the buffer. A standardized protocol was developed for processing urine specimens, including treatment of urine with myoglobin, which abolished the artifactual variation observed previously and enabled reliable electrophoretic separation of the urine RNase isozymes. In addion, myoglobin treatment had a similar positive effect on the electrophoretic behavior of RNase enzymes from some other sources.
Analysis of bacteriophage T4 coat proteins by two‐dimensional electrophoresis and computerized densitometry
Authors: Gersten Dogulas M. , Kurian Ponnamma , Ledly Gary , Park Chan M. , Suhocki Paul V. 
Published: May 2005

AbstractWe have designed treatment conditions which allow analysis of the coat proteins of bacteriophage T4D+ by high resolution two‐dimensional electrophoresis. The best results were obtained when the mature phage particles were subjected to hypertoniclysis and DNase treatment before solubilization with sodium dodecyl sulfate (SDS). Samples containing 250 ug protein gave the optimum resolution, producing 45 spots as visualized by Coomassie Blue staining. Computerized densitometry measurements showed that these proteins range in relative abundance from less than 0.22% to 11.8% of the total.
The fabrication of multichannel tubing and its use in a continuous‐flow isielectric focusing apparatus to give smooth laminar flow
Authors: Quarmby Christopher 
Published: May 2005

AbstractDetails are given of a method of casting multichannel tubing in silicone rubber in the form of a slab 2 mm thick and up to 380 mm long. Up to 264 parallel channels were formed at 0.8 mm centers along the length of the material, each channel being 0.4 mm in diameter. The material was used both as the outlet manifold in an apparatus for continuous‐flow isoelectric focusing (IEF), the construction of which is described, and also in conjunction with a slow‐running peristaltic pump as multichannel pump‐tubing when it was found that at a flow‐rate of about 0.3 ml h−1 per channel, the variation between channels was less than 0.6%. The coupling of the pump‐tubing to the outlet manifold of the separation cell of the IEF apparatus resulted in a very smooth laminar flow of liquid in the separation cell itself.
Isolectric patterns of human alpha1‐ antichymotrypsin (A1AChy) and A1AChy‐protease complexes
Authors: Gianazza Elisabetta , Arnaud Philippe 
Published: May 2005

AbstractWhen studied by thin‐layer polyacrylamide gel isoelectric focusing, alpha1‐an‐tichymotrypsin (A1AChy) in human plasma presents a microheterogeneous pattern, consisting of seven bands with isoelectric points (pI) between 4.1 and 4.45. After removal of sialic acids by neuraminidase treatment, four bands are seen with a more alkaline pI (about 1.0 pH unit). Thus, the microheterogeneity of human A1AChy is due only in part to differential sialylation of the isoproteins. Incubation of A1AChy (in excess) with alpha‐chymotrypsin results in the formation of a primary complex. In the presence of excess protease, incubation results in a secondary complex with a lower pI. Upon incubation of A1AChy with trypsin, no protease‐inhibitor complexes are observed, although there is evidence for partial degradation of the A1AChy molecule.
Electrophoresis of adsorbed albumin
Authors: Absolom Darryl R. , Michaeli Isaac , van Oss Carel J. 
Published: May 2005

AbstractIt was recently observed that bovine serum albumin (BSA), adsorbed onto part of the surface of a glass slide, diffuses along the unoccupied part of that surface (without redissolving in the bulk liquid) at a much greater velocity than the normal diffusion rate of BSA, free solution. In this work a study is made of the electrophoretic transphoretic transport of BSA, adsorbed onto part of the surface of glass slide, along the hitherto unoccupied part of the glass surface. The electrophoretic mobility of adsorbed BSA along the unoccupied glass surface proved to be up to ≅ 2 times faster than in electrophoresis on cellulose acetate strips at the same pH and ionic strength, and up to ≅ 1.6 times faster than in moving boundary electorphoresis. The increase in electrophoretic mobility of adsorbed BSA is most pronounced at the highest density of adsorption. At that density the oblong BSA molecules are closely packed, most probably adsorbed in a tight monolayer, with each molecule positioned perpendicular to the glass surface (although other configurations cannot be excluded). The major driving force enhancing the diffusivity, as well as the electrophoretic mobility of adsorbed BSA, appears to be the high surface pressure of he tightly packed BSA layer. A tentative explanation is offered for the low resistance BSA molecules encounter in moving parallel to the glass surface while remaining adsorbed to the liquid, involving free energies of adhesion of individual protein molecules in various orientations.
Determination of molecular weights and Stokes' radii of non‐denatured proteins by polyacrylamide gradient gel electrophoresis. 2. Determination of the size of stable and labile molecular weight variants of enzymes from plant sources
Authors: Rothe Gunter M. , Purkhanbaba Huschang 
Published: May 2005

AbstractUnder certain conditions in polyacrylamide gradient gel electrophoresis (PAGGE), a linear correlation between the logarithm of the size of calibration proteins (log MW or log Rs) and the square root of their migration distance (√D) can be observed; slope and intercept of the calibration curve depend on the duration of electrophoresis; linearity, however, is maintained over a wide range (4‐60 h, 200 V) (Rothe and Purkhanbaba, Electrophoresis 1982, 3, 33–42.) Using this method the reaction of plant isozyme systems penetrating a linear polyacrylamide (PAA) gradient gel was investigated: lactate dehydrogenase (LDH) from potato tubers behaves similarly to animal calibration proteins. The enzyme exhibits 3 subbands with molecular weights of 144 000, 150 000 and 156 000 (± 1.7 %), respectively. The enzyme system “shikimate oxidoreductase” (SORase) behaves significantly different: during PAGGE it splits into 2 major bands (∼60 000 and ∼110 000, respectively) consisting of several subbands. The results of our experiments show that SORase is present in vivo as a complex system. We believe that this complex is partly indentical with the so‐called pre‐chorismate complex.
Electroimmunochemical analysis of amphiphilic proteins and glycolipids stained with Sudan Black‐containing detergent micelles
Authors: Bjerrum Ole J. , Gerlach James H. , Bøg‐Hansen Thorkild C. , Hertz Jesper B. 
Published: May 2005

AbstractMicelles of non‐ionic detergents can be colored through incorporation of the hydrophobic dye Sudan Black. This property can be used to identify amphiphilic antigens in complex mixtures. Thus, crossed immunoelectrophoresis with 0.05 % v/v Triton X‐100 plus 0.002 % w/v Sudan Black shows selective staining of major amphiphilic proteins from human erythrocytes, thrombocytes and serum, from bovine milk fat globules and from yeast plasma membranes. The method may also give information about glycolipid antigens in detergent‐solubilized membrane material, e.g. mannosyl‐diinositolphosphoryl‐ceramide from yeast. In this case, however, the membrane has to be solubilized with detergent plus Sudan Black and analyzed in detergent‐free gels, where a stable micellar structure seems to exist. In a surplus of detergent, the glycolipid antigen may enter new micelles during electrophoresis, reducing the number of epitopes per micelle below two, thus avoiding precipitation. Analysis of purified lipopolysaccharide from Bordetella pertussis followed this pattern, whereas lipopolysaccharides from Pseudomonas aeruginosa and Escherichia coli, although heavily influenced by the presence of Triton X‐100, were precipitated in the presence of a surplus of detergent. The described method is simple, fast and easy to perform. It requires neither extra equipment nor expensive chemicals. Sometimes heavily stained, detergent‐containing precipitates of a non immunological nature may appear, but these can easily be discerned from immunoprecipitates.
Steady‐state stacking in agarose at various pH
Authors: Buzás Zsuzsanna , Chrambach Andreas 
Published: May 2005

AbstractThe applicability of multiphasic buffer systems to electroendosmosis‐free agarose (IsoGel) was tested in order to be able to combine the benefits of agarose gel electrophoresis with those of steady‐state stacking. It was found that buffer systems of a negative polarity function normally over the entire pH range, whereas proteins with positive net charge are retarded at the gel surface in buffer systems with positive polarity. This retardation is not due to a failure to establish a moving boundary in agarose. Rather, it appears due to a negatively charged weakly acidic functional group in agarose which can be removed from agarose by pre‐electrophoresis at 0.1 M ionic strength. After its removal positively charged proteins also stack normally in multiphasic buffer systems of the positive polarity.
Isoelectric focusing pattern of human antithrombin III (AT‐III): Effects of heparin or thrombin on a microheterogeneous form of AT‐III
Authors: Kera Yoshio , Yamasawa Kichihei 
Published: May 2005

AbstractAfter agarose gel isoelectric focusing (AGIEF) and immunofixation, human antithrombin III (AT‐III) in plasma showed a microheterogeneous pattern which consisted of 5 major and 5 minor components with isoelectric points (pI) between pH 4.8 and 5.2. Treatment with neuraminidase reduced this microheterogeneity of AT‐III to 1 major, 3 sub‐major and 2 or 3 minor components with a shift in pI to between pH 5.5 and 5.8. Thus, differential sialylation of the isoproteins partially contributed to the microheterogeneity observed. The addition of heparin to plasma produced broad acidic precipitates with pIs between pH 4.2 and 4.7 on the gel. The same acidic precipitates were also obtained from heparin‐purified AT‐III mixture. The appearance of these precipitates was related to the decreasing in number or in staining intensity of the native AT‐III bands. These precipitates may be complexes of heparin and AT‐III. The addition of human or bovine thrombin to plasma produced several basic minor components, which were similar to those in serum, at a thrombin concentration of 48 or more NIH U/ml‐plasma. The basic minor components were not obtained from a purified AT‐III‐thrombin mixture. These might be complexes between AT‐III, thrombin and some of the other coagulation factors.
Comparative analysis of subcellular proteins by SDS‐polyacrylamide gel electrophoresis with dorsolateral and ventral prostates of rats
Authors: Matuo Yuhsi , Nishi Nozomu , Negi Tetsuro , Wada Fumio 
Published: May 2005

AbstractProteins in the cytosol, post‐nuclear particulate and nuclear fractions from dorsolateral and ventral prostates of rats were analyzed and compared by sodium dodecyl sulfate (SDS)‐polyacrylamide gel electrophoresis. Protein species separated in the gel were stained with Coomassie Brilliant Blue R‐250 (CBB), and expressed as (apparent molecular weight of the respective species x 10−3)K. (i) In the cytosol fraction the contents of 30K, 67K, 110K, and 120K proteins were abundant in the dorsolateral prostate. On the other hand, 14K and 16K proteins were specific to the ventral prostate. By carbohydrate staining, major glycoproteins were found to be 120K and 16K for the dorsolateral and ventral prostates, respectively. (ii) In post‐nuclear particulate fractions in both lobes, proteins having molecular weights greater than 42 000 were abundant. Of these, the contens of 65K, 105K and 120K were higher in the dorsolateral prostate than in the ventral prostate. Glycoproteins having the same apparent molecular weight as detected in the cytosol fractions were also found in the post‐nuclear particulate fractions of both lobes. (iii) In the nuclear fraction H2A, H2B, H3 and H4 histones were fundamentally similar between the two lobes, whereas H1 histones including two subspecies were slightly but significantly higher in the ventral prostate than the dorsolateral prostate. The most remarkable difference between the lobes was the content of 20K in the dorsolateral prostate which was at least 20 times higher than that found in the ventral prostate.
Isolation of urinary β2‐microglobulin by isoelectric focusing: Use of a new method of protein localization
Authors: Bonitati James , Sabatino Bruce , Van Liew Judith B. 
Published: May 2005

AbstractAn indirect fluorescence method for locating colorless protein bands in focused Sephadex gel slabs, demonstrated previously using standard protein solutions [1, 2], has been applied to a complex natural protein mixture: preconcentrated human urine. Protein bands separated by distances of 2 mm or less in the gel slab were accurately located and excised without cross‐contamination. Urinary pigment did not interfere with the localization technic. Microgram quantities of six protein bands were excised from thin gel slabs, and then partially characterized by cellulose acetate electrophoresis. In addition, the bands belonging to two homologues of β2‐microglobulin (B2M I and II) were identified immunologically after systematic excision of the various protein bands in the focused pattern. This one‐step protein isolation procedure was scaled up to the milligram level for the isolation of B2M‐II. In two experiments, 1.60 mg and 2.22 mg were isolated in 73 % and 78 % purity, with yields of approximately 55 % and 76 %.
Analysis of the genetic variants of the human Gc system (VDBP) by isoelectric focusing in immobilized pH gradients
Authors: Cleve Hartwig , Patutschnick Walburgis , Postel Wilhelm , Weser Johann , Görg Angelika 
Published: May 2005

AbstractThe genetic variability of the human group‐specific component (Gc) system has been examined by isoelectric focusing with immobilized pH gradients. The phenotypes of the six common Gc types and of 15 rare genetic Gc variants are described and presented. High resolution of the Gc components is accomplished. The classification can be made without specific immune reaction. Isoelectric focusing with immobilized pH gradients represents a powerful tool for analyzing genetic variability.
Description and detection of moving sample zones in zone electrophoresis: Zone spreading due to the sample as a necessary discontinuous element
Authors: Thormann Wolfgang 
Published: May 2005

AbstractThis paper deals with the description and detection of moving sample zones in free zone electrophoresis. It is shown that the shape of a sample zone never reaches a steady‐state. However, either the front or the rear boundary of the sample zones, but not both, can represent steady‐state concentration distributions with decreasing peak heights. This results in an unsymmetrical broadening of the moving zones. With a multichannel detection device, consisting of 255 equidistant detectors along the separation trough, asymmetric electric field distributions of the sample components are measured. These are shown to agree with theoretical predictions, made by a model based on migration only. This clarifies the fact that in zone electrophoresis migrational dispersion due to the applied electric field is usually much larger than diffusional spreading.
Staining of Concanavalin A‐reactive glycoproteins on polyacrylamide gels with horseradish peroxidase — a critical evaluation
Authors: Schott Klaus‐Joachim , Neuhoff Volker , Nessel Birgit , Pötter Ulla , Schröter Joachim 
Published: May 2005

AbstractThe method to localize and quantitate glycoproteins on polyacrylamide gels is based on the binding of Concanavalin A to the glycoproteins and the subsequent coupling of horseradish peroxidase to the glycoprotein ‐ Concanavalin A complex. The complex is visualized by the oxidation of 3,3 ‐diaminobenzidine with horseradish perexidase ‐ H2O2, forming a brownish stain at the gel surface. Various parameters are analyzed and a scheme for the optimal staining of macro and micro slab gels is given. The quantitative evaluation of the stained gels is demonstrated. The staining and quantitation of nitrocellulose transfers is also shown.
A photodevelopment silver stain for the rapid visualization of proteins separated on polyacrylamide gels
Authors: Merril Carl R. , Harrington Michael , Alley Valerie 
Published: May 2005

AbstractBy using light (“photodevelopment”) to develop a metallic silver image of proteins separated on polyacrylamide gels, it is possible to visualize protein and nucleic acid patterns within 10 min after electrophoretic separation. This light catalyzed silver stain is based on establishing gel conditions in which ionic silver is reduced to silver metal in the presence of light energy at a different rate in gel regions containing biopolymers (such as proteins or nucleic acids) than the rate of reduction in regions which are devoid of such biopolymers. This photodevelopment method requires only two solutions: a solution to “fix” the proteins and a solution containing silver ions, which produces an image when exposed to light. This type of protein stain has achieved a sensitivity of 0.5 ng of protein. DNA separated on polyacrylamide may also be visualized with this stain. However, DNA generally produces a negative image. Studies of photodevelopment silver stains allow further insight into the processes which are involved in the ability of silver to form images of biopolymers separated by electrophoresis.
Bidirectional analytical isotachophoresis: Simultaneous determination of anionic and cationic boundaries
Authors: Thormann Wolfgang , Arn Dieter , Schumacher Ernst 
Published: May 2005

AbstractThis paper deals with electrolyte systems capable of producing suitable isotachophoretic zone structures in either migration direction. These systems contain moving reaction boundaries. A dilute solution of a strong acid as anolyte and the sodium salt of a weak acid as catholyte are analyzed to provide insight into the features of such dual systems. For various electrolyte arrangements the boundaries are examined for resolving capability in isotachophoresis. Both anionic and cationic boundaries were followed simultaneously using a capillary type apparatus with an array detector.