摘要:

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

ISSN:0173-0835    

DOI:10.1002/elps.1150030202

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1982

数据来源: WILEY

摘要:

AbstractTwo potentialobstacles to using agarose gel electrophoresis for the fractionation of viruses and related particles are: (a) adherence of the sample to the gel during electrophoresis, and (b) the prolonged procedures necessary for determining whether or not particles stained with nucleic acid‐specific stains have their nucleic acid in a packaged state. It is demonstrated that adherence of positively charged tail fibers of bacteriophage T7 to agarose gels is the probable cause of a previously described (Serwer, P. and Pichler, M. E.,J. Virol.1978,28, 917–928) sample concentration‐dependent spreading of bands formed by bacteriophage T7 during agarose gel electrophoresis (concentration effect). Procedures for screening preparations of agarose for nonadherence of particles are described and use of these procedures has resulted in the finding that the concentration effect is decreased or eliminated for T7 in preparations of agarose designed for low electro‐osmosis. In addition, it has been shown that after electrophoresis, the intensity with which bacteriophage T7 (and other DNA bacteriophages) stains with the nucleic acid‐specific stain, ethidium bromide, is increased by ejecting DNA from the interior of T7; this increase in staining intensity is used to recognized bands formed by particles with pac

ISSN:0173-0835    

DOI:10.1002/elps.1150030203

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1982

数据来源: WILEY

摘要:

AbstractBacteriophage T4, bacteriophage T4 without its tail fibers, and bacteriophage P22 without its tail have been subjected to electrophoresis in agarose gels; electrophoretic mobility (μ) of these particles as a function of the concentration of agarose (A) has been determined using preparations of agarose with different amounts of electro‐osmosis. When μ was extrapolated to anAof 0 (μo), it was found that a term, μE, constant for each preparation of agarose must be added to μoto correct for electro‐osmotic flow. A relationship between μEand a manufacturer‐determined coefficient of electro‐osmosis has been empirically derived. Corrected μo's are in agreement with data obtained by solid support‐free

ISSN:0173-0835    

DOI:10.1002/elps.1150030204

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1982

数据来源: WILEY

摘要:

AbstractA rapid preparative isoelectric focusing agarose slab gel technique that should prove to be valuable for the purification and characterization of clinically important macromolecules is described in this report. Up to 120 mg of plasma proteins have been focused in a 9.5 ml volume IsoGel agarose slab gel on a hydrophilic polyester plastic backing (GElBond) in 30 min or less. The agarose was permeable to proteins with molecular mass of 5–50 × 106daltons and allowed recoveries of from 11–100 % depending on the proteins electrofocused. Proteins whose isoelectric points (pI) differed by 0.07 pH units will be separated by the described techn

ISSN:0173-0835    

DOI:10.1002/elps.1150030205

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1982

数据来源: WILEY

摘要:

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 fromBordetella pertussisfollowed this pattern, whereas lipopolysaccharides fromPseudomonas aeruginosaandEscherichia 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 imm

ISSN:0173-0835    

DOI:10.1002/elps.1150030206

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1982

数据来源: WILEY

摘要:

AbstractLine immunoelectrophoresis was adapted to the quantitative analysis of antigens directly from tissues without any previous extraction. The technique was used in a study on the production and the distribution of two α‐amylase isozymes (I and II) in tissues of wheat seeds that sprouted (radicle emergence) after different periods of germination. The α‐amylase content was examined in the scutellum and in three tissue fractions dissected from a 0.5 mm slice parallel and next to the scutellum (aleurone layer, outer and inner part of starchy endosperm). At the stages of germination investigated, α‐amylase I was not detected in these tissues. The results indicate that α‐amylase II accumulated in the scutellum and also in the aleurone layer after one day's germination; after about two day's germination, the α‐amylase II content in the aleurone layer was similar to or higher than that in the scutellum. Moreover, the production and distribution of α‐amylase II do not seem to be correlated with the

ISSN:0173-0835    

DOI:10.1002/elps.1150030207

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1982

数据来源: WILEY

摘要:

AbstractSeparation of ATP and ADP was achieved in isotachophoresis by using malonate, inorganic phosphate (Pi), creatine phosphate and lactate as spacers, which isolate these nucleotides from other UV‐absorbing products in rat tissues. By combining isotachophoresis with enzymatic end‐point analysis of several rat tissues, calibration for ATP and ADP was achieved obtaining an operative calibration curve that allows the use of isotachophoresis as the only analytical technique in further assays. ATP and ADP quantities of rat skeletal and smooth muscle, heart, liver, kidney and lung were determined by isotachophoresis. Individual variation was also calculated by analyzing 30 animals in the same control conditions, muscular tissues showing the greatest values for ATP and ADP, whereas kidney and lung show the smaller values. ATP/ADP ratio is discussed as a representative parameter for describing the energy content in biological tissues, there are also experimental reasons which imply that other product concentrations in this parameter not be included. This ATP/ADP ratio is calculated in the rat tissues studied, as well as its individual variation, obtaining the larger values for muscular tissues and the smaller for kidney. A great individual variation is found in kidney, followed by heart and skeletal muscle, and the most constant values are found in liver and smooth muscle. The results obtained in this calibration method for isotachophoresis assay suggest the use of a similar procedure in order to apply this technique for other nucleotide analy

ISSN:0173-0835    

DOI:10.1002/elps.1150030208

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1982

数据来源: WILEY

摘要:

AbstractPepsin‐solubilized type II collagen and some of its CNBr‐derived peptides could be stained for carbohydrate with periodic acid‐Schiff (PAS) after polyacrylamide gel electrophoresis. PAS intensely stained an aldehyde‐containing peptide (Mr>10 000) which was not detected with Coomassie Brilliant Blue R‐250 stain. Types I and III collagens and their CNBr peptides stained poorly, if at all,

ISSN:0173-0835    

DOI:10.1002/elps.1150030209

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1982

数据来源: WILEY

摘要:

AbstractThe electrophoretic mobility of a number of microspheres of homo‐ or copolymerized acrolein has been studied. It was found that increasing amounts of methacrylic acid increased the negative mobility of the microspheres, whereas reaction of the microspheres with 1,6‐diaminohexane decreased the negative mobility. When coated with antibody and specifically bound to cells, these microspheres were found to alter the target cells' electrophoretic mobility. This process allowed the separation of synthetic mixtures of microsphere‐labeled and unlabeled cell populations. Over 90 % of the recovered cells were found in fractions that were essentially either pure labeled or unlabeled

ISSN:0173-0835    

DOI:10.1002/elps.1150030210

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1982

数据来源: WILEY

ISSN:0173-0835    

DOI:10.1002/elps.1150030211

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1982

数据来源: WILEY