bluuebunny/crossref_metadata_2025_split huggingface dataset:
pH‐Gradient development and focusing speed in thin‐layer polyacrylamide gel isoelectric focusing: A comparison between PharmalyteR, AmpholineR and ServalytR
Authors: Låås Torgny , Olsson Ingmar 
Published: May 2005

AbstractThe rate of pH gradient development and protein migration towards their isoelectric points(pI) were investigated for thin‐layer polyacrylamide gel isoelectric focusing (PAGIF) using Pharmalyte 3–10, Ampholine 3.5–9.5 and Servalyt 2–11 as carrier ampholytes. Using volt‐hours(Vh) as the controlling parameter, it was found that both pH gradient development and protein focusing proceeded very similarly with all three carrier ampholytes, Only two significant (but minor) differences between the carrier ampholytes were observed: (1) due to a faster voltage rise with Pharmalyte, the Vh needed for focusing are achieved faster than with the other two carrier ampholytes and (2) focusing of protein with alkaline pI's (> pH 8) is slightly slower with Servalyt (both in time and Vh). This is presumably explained by the low field strength at the alkaline end of the pH gradient with Servalyt. Also, it was found that pH gradient development was completely independent of applied voltage when Vh were used as controlling parameter.
Polymerization kinetics of polyacrylamide gels. III. Effect of catalysts
Authors: Righetti Pier Giorgio , Gelfi Cecilia , Bosisio Adriana Bianchi 
Published: May 2005

AbstractThe kinetics of photopolymerization, in presence of riboflavin or riboflavin‐5′‐phosphate (FMN), have been studied either spectrophotometrically, at 283 nm, or chemically, by titrating unreacted double bonds with permanganate. The two sets of data are in good agreement and suggest that at least 8 h of light exposure are needed to ensure 95% conversion of monomers into polymers. Up to now, it was generally accepted that photopolymerization should proceed for 1 h. It is not necessary to prolong light exposure for longer than 8 h, because even a 24 h illuminatin period only improves the conversion from 95% to 96%. Although conditions are described which ensure at least 95% conversion of both riboflavin‐and persulfate‐catalyzed gels, their visco‐elastic properties are widely different, suggesting that the final structure of these two matrices must vary considerably as a function of the catalyst used.
Comparison of rare PGM1 variants by isoelectric focusing and conventional electrophoresis: Identification of five new variants
Authors: Dykes Dale D. , Polesky Herbert F. 
Published: May 2005

AbstractA comparison of specimens tested in our laboratory for the red cell enzymes sub‐system PGM1 by isoelectic focusing and conventional electrophoriesis demonstrated 11 rare allotypes. In addition to the allotypes PGM, PGM, PGM, , and , we indengified five new variants: , , , , . Of the five new variants only and could be separated from common PGM1 variants on conventional elehoresis. Because of the unique banding patterns for the variants observed, it was found necessary to use both isoelectic focusing and conventional methods for proper identification. Due to the presence for several different momeclatures for naming PGM variants, it is felt that a universal nomenclature should be defined to deal with PGM variants using both methods of analysis.
Synthetic inorganic ion exchangers XXI. Electrochromatographic separations of metal ions on lanthanum antimonite‐impregnated paper
Authors: De Anil K. , Chakraborty Punyabrata 
Published: May 2005

AbstractElectrochromatographic studies of several metal ions on lanthanum antimonateimpregnated papers have been carried out. Various background electrolytes were used for these studies at different voltage and time intervals. On the basis of the differential mobilities of metal ions, which depend on the ion exchange properties of lanthanum antimonite and the nature of complex formation with the electrolytes, some important binary and ternary separations have been achieved.
A technique for the detection of variable electrophoretic patterns of hair proteins
Authors: Budowle Bruce , Acton Ronald T. 
Published: May 2005

AbstractA reproducible isoelectic focusing method has been developed which displays the structural proteins of hair. By staining with the photochemically derived stain, hair protein variability was observed. On a limited number of individuals, five different isoelectric focusing patterns were detected. In addition, polychromatic differences of proteins with similar isoelectric points were found.
Silver staining of DNA in polyacrylamide gels: Linearity and effect of fragment size
Authors: Goldman David , Merril Carl R. 
Published: May 2005

AbstractA simplified, inexpensive silver stain for proteins in polyacrylamide gels also stains DNA with a sensitivity of 1 ng (< 0.03 ng/mm2). Sensitivity of detection is superior to ethidium bromide and, for 11 double‐stranded fragments of ϕX174 DNA, staining is linear with respect to concentration. Surprisingly, the slope of staining is similar for pieces of DNA larger than 300 base pairs, but is greater for fragments of smaller size. Staining of DNA with silver is a practical alternative to visualization with ethidium bromide and may serve as a tool for elucidating the basis of silver staining.
Rapid staining of proteins in ultrathin‐layer isoelectric focusing in polyacrylamide gels
Authors: Frey Manuela D. , Radola Bertold J. 
Published: May 2005

AbstractA method for protein staining after isoelectric focusing (IEF) in polyacrylamide gel is described in which the total time for fixation, staining and complete gel background destaining is reduced to 3–5 min. This is achieved by (i) staining dry gels on silanized supports instead of hydrated gels, (ii) reducing gel thickness to 50 μm, (iii) using Servalyt carrier ampholytes with good destaining properties, and (iv) using dyes with low affinity for carrier ampholytes, namely Serva Violet 17 or 49, facilitating destaining of gel background. The sensitivity of staining is 100–200 ng protein/ mm2over a 10 cm separation distance and 20–30 ng protein/ mm2over 3 cm gels (miniature focusing). Addition of 5–20 % methyl acetate to the staining or destaining solution reduces the sensitivity of staining significantly. Most effective fixation of proteins in 50 μm gels is achieved with 20% trichloroacetic acid. In solutions of alcohol and acetic acid, appreciable amounts of proteins are leached after incubation for 5–20 min. Paraformaldehyde, malondialdehyde or glutaraldehyde (5–10 %) exhibit no fixative effect on proteins in 50 μm gels when used directly or after precipitation of the proteins with trichloroacetic acid. The selective leaching of proteins under a variety of experimental conditions shows that adequate fixation is particularly important in work with basic, low molecular weight and/or hydrophobic proteins.
Gel electrofocusing in a natural pH gradient of pH 3–10 generated by a 47‐component buffer mixture
Authors: Cuono Charles B. , Chapo Giselle A. 
Published: May 2005

AbstractA natural pH gradient with a pH range of 3–10 was generated from a mixture of 47 buffers utilizing a polyacrylamide gel slab format. The gradient is stable for at least 18 h at 100 V/cm of gel, with the attainment of the steady state at 1 h and of isoelectric protein positions within 2.5 h. Gels containing buffer constituents at a final concentration of 10–20 mM exhibit a conductivity which is approximately one order of magnitude less than that exhibited by any synthetic carrier ampholyte mixture (SCAM), thereby permitting a high degree of resolution in analytic applications. Excessive Joule heating and artefactual protein‐protein interaction, both potential drawbacks to electrofocusing at lower ionic strengths, were not observed in this study. The pH gradient generated by the buffer mixture makes possible for the first time electrofocusing of proteins under constant conditions in longitudinal studies, independently of the source and batch of carrier constituents. Furthermore, this system completely circumvents the physical, chemical and biologic problems frequently encountered with SCAMs.
Specific antiserum staining of two‐dimensional electrophoretic patterns of human plasma proteins immobilized on nitrocellulose
Authors: Anderson N. Leigh , Nance Sharron L. , Pearson Terry W. , Anderson Norman G. 
Published: May 2005

AbstractHuman plasma proteins separated by high‐resolution two‐dimensional electrophoresis have been electrophoretically transferred to sheets of nitrocellulose using a modification of the method of Towbin, Staehelin, and Gordon [8]. Although the proteins have been denatured in sodium dodecyl sulfate and separated into subunits, the nitrocellulose‐bound molecules still react with appropriate specific antisera even after storage of the transfer in air at room temperature for 5 months. Of 25 proteins whose location in the pattern had been previously determined, 24 are specifically revealed on transfers of whole plasma patterns by appropriate antiserum. In addition, 6 previously unidentified proteins (prothrombin, C1s, C4γ, C1s inhibitor, Ig J‐chain, and a1AP glycoprotein) have been identified in the pattern for the first time using the transfer technique. It therefore seems likely that a large majority of proteins (> 96 % in this study) retain sufficient conformation throughout the analytical procedure (or can regain it easily afterwards) to be recognized immunologically. The transfer technique thus constitutes a generally useful immunological “third‐dimension” in the high‐resolution separation of proteins. Of three monoclonal antibodies similarly tested, none could detect antigen transferred to nitrocellulose from a two‐dimensional gel, while each bound specifically to the appropriate antigen absorbed in native form to the nitrocellulose.
Improvements of isoelectric focusing in agarose for direct tissue isoelectric focusing
Authors: Thompson Basil J. , Dunn Michael J. , Burghes Arthur H. M. , Dubowitz V. 
Published: May 2005

AbstractImprovements of isoelectric focusing (IEF) in agarose are described that result in consistently obtaining broad, linear pH gradients in the range pH 4 to 9.2 at equilibrium. Cathodic drift has been reduced by the use of a blend of commercial carrier ampholytes and by incorporating lysine and arginine in the catholyte as spacers. Resolution has also been enhanced by the use of thin gels, supplemented with sorbitol and glycerol. In addition, a comparison of two agarose preparations for isoelectric focusing was performed using two commercial carrier ampholytes. To minimise background staining, Triton complexes were removed by modification of the fixing and clearing procedures. These modifications to IEF in agarose are being applied to direct tissue isoelectric focusing (DTIF) of normal and diseased human skeletal muscle.
Detection of bacteriophage‐antibody complexes by agarose gel electrophoresis
Authors: Serwer Philip , Hayes Shirley J. 
Published: May 2005

AbstractTo detect bacteriophage‐antibody complexes, specific antiserum has been incubated with bacteriophages and antiserum‐induced aggregation has been detected by agarose gel electrophoresis, using differential sieving of the gel to fractionate aggregates by size. Agarose gel electrophoresis should also be of use in monitoring reaction of other virus‐sized antigens with antibodies.
Design types and performances of preparative polyacrylamide gel electrophoresis apparatus
Authors: Akaiwa Shigeo 
Published: May 2005

AbstractThe designs of both gel and elution chambers in preparative electrophoresis apparatuses are key elements for their performance. The present paper describes four types of designs which were developed by the author to alleviate the shortcomings found in the preceding designs. Performance tests, using human hemoglobin (Hb), partially purified haptoglobin 2‐1 type (Hp 2‐1), and bis‐albumin‐containing serum, show that insufficient dissipation of Joule heat and a heterogeneous electric field cause curvature and slant of the sample zones, and that only a chamber of optimal shape and size leads to minimal dilution, giving a good resolution. Moreover, notch, angle, and rim (Fig. 5), designed to support the gel, and tapering of the elution chamber to minimize its volume, are also important ministructures for the performance of the instrument. Unoptimal sets of these structures also cause concave sample bands and result in bad resolution and excessive dilution. These findings emphasize that in scaling up the apparatus, the structure must be designed on the basis of the detailed fundamental experiments from macrostructure to microstructure by using models of actual size.
Implementation and application of a method to quantitate 2‐D gel electrophoresis patterns
Authors: Jansson Peter A. , Grim Larry B. , Elias John G. , Bagley Elisabeth A. , Lonberg‐Holm Karl K. 
Published: May 2005

AbstractWe describe a system for accurate photometry and analysis of two‐dimensional (2‐D) gels, and its application to both high and low resolution gels. Image acquisition is video‐based and includes methods of data correction necessary for accurate photometry. The analysis is based upon a method described previously by Lutin et al. [8], which we have modified for improved stability and adapted to run on our PDP 11/60 computer. We illustrate analysis of a typical gel with a sequence of images showing the data while Gaussian components are estimated, removed, added and fitted as the iterative algorithm proceeds. We conclude by demonstrating the use of a mathematical curve‐of‐growth model for calibrating peak volumes vs. protein loading.
An improved electroelution method for the recovery of DNA fragments from agarose and acrylamide gels
Authors: Ho Nancy W. Y. 
Published: May 2005

AbstractA simple elution tube has been constructed which allows reliable electroelution of DNA restriction fragments from agarose and acrylamide gels. This method enables the recovery of DNA fragments of 50 to over 50 000 base pairs from cylindrical or slab gels with yields between 70 % to 90 %. This method has several distinct features which make it outstanding. (1) It concentrates DNA from a large sample of gels into a small volume of buffer (less than 500 μl) during elution oso that the recovered DNA can be directly precipitated out with alcohol for subsequent use. (2) The recovered DNA is contaminated with only a minimal amount of agarose or acrylamide gel so that it may be subsequently digested with most restriction endonucleases with normal efficiencies. (3) It is particularly useful for the recovery of DNA fragments that have been subjected to end labeling with [γ‐32P] ATP and polynucleotide kinase, because it not only recovers the DNA from the gel, but also purfies the fragments from the contaminating background [γ‐32P] ATP. (4) Many different sizes of DNA fragments can be recovered at the same time. In addition, the elution tubes are easy to construct with not much more cost than that of a piece of plexiglas tubing.
Studies to establish a reference pattern of slime‐forming, encapsulated Streptococcus cremoris plasma membrane antigens by crossed immunoelectrophoresis
Authors: Niskasaari Kaarina , Valkonen Kaija H. , Forsén Raili 
Published: May 2005

AbstractThe antigenic composition of the plasma membrane of the slime‐forming, encapsulated strain Streptococcus cremoris T5, originally isolated from the Finnish ropy sour milk product “viili”, was studied by crossed immunoelectrophoresis (CIE). Membrane proteins were solubilized with 1% Triton X‐100 extraction and mild sonication at pH 8.7. The solubilizing effect for membrane protein was investigated by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE). Only quantitative differences were observed in the SDS‐PAGE patterns of the Triton X‐100 extracts as compared with that of the total‐protein sample. The anti‐membrane antiserum pool was collected over twelve months. The conditions for CIE were standardized using different antigen and antibody concentrations. The immunoprecipitation pattern with standard antisera was reproducible with separate membrane preparations. The solubilized membrane antigens could be stored at ‐20°C for two months with no changes in the precipitation pattern. A reference immunoprecipitation pattern with 19 precipitation lines was established for S. cremoris T5 membrane antigens. Two immunoprecipitates had a strong staining intensity with Coomassie Brilliant Blue, and one of these was also stainable with periodate‐Schiff's reagent. Certain special characteristics of these precipitation lines are proposed for further study.
Detection of DNA and lipopolysaccharide in thick polyacrylamide gels using an improved silver stain
Authors: Marshall Thomas 
Published: May 2005

AbstractA methylamine‐incorporating silver stain is described for the detection of deoxyribonucleic acid (DNA), protein and lipopolysaccharide in thick (3 mm) polyacrylamide gradient gels (4–20 % w/v). Its sensitivity for DNA and protein is more than 100 times greater than ethidium bromide and Coomassie Brilliant Blue R‐250, respectively. It detects 0.01 ng DNA, 0.16 ng of human serum albumin and ng quantities of lipopolysaccharide. DNA gives red bands whereas protein and lipopolysaccharide give brown/black bands. The staining procedure is inexpensive, highly reproducible and overcomes the problem of background stain associated with thick gels. It also gives excellent results with thin (1 mm) gels and can be used to stain a number of gels simultaneously. A simplified staining procedure is also discussed.
An electrophoretic investigation of plasma fibronectin by immunofixation after agarose gel electrophoresis
Authors: Castle Stephen L. , Board Philip G. 
Published: May 2005

AbstractAn electrophoretic technique for the rapid screening of fibronectin from plasma and serum is described. This technique was used to screen nearly 3000 human plasma and serum samples from different racial groups as well as plasma samples from dogs, cats, pigs, goats, mice, donkeys, sheep, horses, camels, cattle and Highland ponies for genetic variants. Only one possible genetic variant, from an Indonesian population, was detected. Other potential uses of the electrophoretic technique are discussed.
Placental affinity for pregnancy‐associated murine protein‐1 examined by crossed affino immuno‐electrophoresis
Authors: Hau Jann , Teisner Bøsrge , Svendsen Per 
Published: May 2005

AbstractUsing crossed affino immunoelectrophoresis and tissue extracts added to the first‐dimension gels the affinity between pregnancy‐associated murine protein‐1 (PAMP‐1) and various tissues was examined. Substances binding to PAMP‐1 in the liver and in the placenta were found to affect the electrophoretic migration velocity of PAMP‐1 when added to first‐dimension gels.
Electrophoresis and recovery of active mRNA from composite ultra low/medium gelling temperature agarose gels
Authors: Fourcroy Pierre 
Published: May 2005

AbstractThe electrophoretic separation of RNA species in composite ultra low/medium gelling temperature agarose is described. The resolution obtained was superior to that observed with the standard medium gelling temperature agarose alone. Poly(A)‐containing mRNA was successfully fractionated and recovered in a biologically active form from the separating media.
Changes of the electrophoretic mobility of human monocytes are regulated by lymphocytes
Authors: Bauer Johann , Hannig Kurt 
Published: May 2005

AbstractThe changes of the electrophoretic mobility (EM) of human monocytes during invitro maturation into macrophages were investigated. Incubation of isolated monocytes for longer than 4 days was always accompanied by an increase of the EM of the monocytes. This increase of the EM could only be inhibited when lymphocytes were incubated together with the monocytes. The inhibitory effect of the lymphocytes was optimal when the culture medium was supplemented by autologous plasma and when the cell suspension was kept in teflon beakers, but it was reduced when the culture medium was supplemented by fetal calf serum (FCS) and when the cell suspension was kept in polystyrene dishes. The macrophages, which showed increased EM after 10 days of incubation, had equal antibody‐dependent cell‐mediated cytotoxic (ADCC) activity but lower plaque‐forming activity than the macrophages which retained their original EM during incubation. The finding of macrophage sub‐populations which differ in their EM provides a good opportunity to isolate and further investigate macrophages, which may have different biological activities.
Gel electrofocusing in tubes containing Sephadex
Authors: der Lan Birgit An , Ben‐or Sarah , Allenmark Stig , Sullivan James V. , Fitze Paul , Jackiw Andrew 
Published: May 2005

AbstractGel electrofocusing (EF) on Sephadex G‐75, which provides relatively non‐restrictive pore sizes and relatively broad protein concentration distributions (which reduces concentration‐dependent aggregation), was adapted for use in glass tubes. This allows for EF with meticulously controlled low temperatures in conjunction with large sample and gel volumes. Specifically, the technique was developed for EF of steroid receptors, which are highly sensitive to temperature (temperature transformed), and whose aggregation behavior excluded the use of polyacrylamide or agarose gels. The technique uses the temperature‐controlled apparatus designed for quantitative polyacrylamide gel electrophoresis, with three additions. These are a) a 12‐place evacuation device for forming simultaneously 12 cylindrical Sephadex gels (either 6 or 18 mm diameter), b) an electrode arrangement capable of maintaining the same regulated voltage across several gel tubes, and c) a jacket and lid of polyurethane foam to insulate the electrolyte chambers of the EF apparatus.
Dansylated proteins as internal standards in two‐dimensional electrophoresis and Western blot analysis
Authors: Lubit Beverly W. 
Published: May 2005

AbstractDansylated proteins have been used in combination as isoelectric point and molecular weight standards and included in gel samples for separation by two‐dimensional polyacrylamide gel electrophoresis. These proteins provide invariant points of reference that facilitate the unequivocal identification of proteins separated by two dimensional polyacrylamide gel electrophoresis without affecting resolution. Moreover, immunoautoradiograms of Western blots can be aligned with their corresponding gel patterns with great accuracy. The logical extension of the established usage of dansylated protein standards to these applications is described.
A novel human kidney‐specific protein detected by two‐dimensional electrophoresis: Isolation, radioimmunoassay, and immunohistochemical localization
Authors: Ziai Mohammed Reza , Jackson Peter , Thompson R. J. 
Published: May 2005

AbstractHigh resolution two‐dimensional polyacrylamide gel electrophoresis has been used to detect a soluble protein in extracts of human kidney which was not present in the electrophoretograms of other human tissues. This protein, coded kidney‐acidic protein‐60 (KAP‐60) was purified to homogeneity and was characterized as having a native molecular weight of 85000, a monomer molecular weight of 35 000 and an isoelectric point of 5.6. KAP‐60 was devoid of detectable carbohydrate or phosphate groups. Injection of the protein into rabbits produced antisera which were used to develop a radioimmunoassay for the protein and to localize the protein to specific cells within the kidney by the immunoperoxidase technique. A survey of human tissues by radioimmunoassay showed that KAP‐60 was present in human kidney at approximately 150 times the concentration found in human liver and 300 times the level found in other human organs examined. Immunohistochemistry localized the protein to the proximal and collecting tubules in the kidney with no staining of glomeruli or distal tubules. The present study illustrates the use of high‐resolution two‐dimensional electrophoretic techniques in the detection of human tissue‐specific proteins which may prove of clinical value in the biochemical diagnosis of tissue‐specific damage.