摘要:

AbstractAgarose gel electrophoresis has been shown to give rise to non‐linear plots of log (mobility)vs.gel concentration of spherical viruses (Serwer,[1]) and cellular vesicles (Gottliebet al.[2]). This finding also applies to proteins as shown in this study. Considering that in the non‐linear plot, the slope becomes a function of gel concentration, it is possible to determine particle properties and gel parameters by a modification of the conventional method derived from the Ogston theory for long‐fiber gels. This treatment shows: (a) In application to data obtained from gel electrophoresis (0.4 to 1.6 % agarose) of viruses (13 to 42 nm radius) and with increasing gel concentration: (i) an increase of apparent total fiber length per g agarose matrix, (ii) a reduction of apparent fiber radius and (iii) a constant fiber volume (per g matrix material) of the agarose fiber. The values of the fiber radii identify the fibers as agarose supercoils (or aggregates of it) with a radius of 20–55 nm. (b) In application to data obtained from gel electrophoresis (1.2 to 8 % agarose) of proteins (1.7 to 5.8 nm radius): a fiber volume indicative of additional sieving by the agarose double helix (of known radius of 0.5–2 nm). This is in agreement with a previous suggestion by Serwer [1]that proteins are able to penetrate into the double‐helical network of which the agarose supercoil consists. (c) The likelihood of a continuous transition from case (a)

ISSN:0173-0835    

DOI:10.1002/elps.1150070602

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

年代:1986

数据来源: WILEY

摘要:

AbstractRecently, we described hybrid isoelectric focusing in rehydrated polyacrylamide gels as an analytical procedure which is based on separation in immobilized pH gradients supplemented with low concentrations of carrier ampholytes (Altland and Rossmann,Electrophoresis1985,6, 314–325). When using this procedure at various pH ranges, in the presence of reducing agents and carrier ampholytes at different concentrations and from different suppliers, over prolonged time intervals for separation and at various acrylamide concentrations in the gel, liquid exudation has been observed on the gel surface which may result in a confluent liquid layer adversely affecting the final separation pattern. The effect is demonstrated and procedures are described to overcome the phenomenon by adding polyols like glycerol, sorbitol, sucrose or dextrans to the rehydration solution. Comparable effects are achieved when these components are added at the same concentration by weight rather than by moles. The addition of 10 g % Dextran 8 or 20 g % sucrose or sorbitol to the rehydration solution prevents liquid exudation for several hours. These additives are compatible with high concentrations of urea, with reducing agents and with neutral detergent (Triton X‐1

ISSN:0173-0835    

DOI:10.1002/elps.1150070603

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

年代:1986

数据来源: WILEY

摘要:

AbstractImmobilized pH gradient (IPG) slab gels are conventionally formed by use of two‐chamber gradient mixers on pH‐neutralized Immobiline mixtures, co‐polymerization of the gradient of Immobilines with acrylamide at 50 °C, washing and drying of the gel to its original weight followed by pre‐electrophoresis before applying the sample. This tedious procedure was replaced by one using gradient formation by pump, eliminating pH neutralization of the monomer mixture, substituting polymerization at 50 °C for one at the temperature of electrophoresis (10 °C in this study) and omitting washing and drying of the gel prior to use. Carrier ampholyte (CA) containing IPG gels were formed in the same way except that CAs were added to the polymerization mixture. Pre‐electrophoresis of IPG gels, formed by either the conventional or the simplified procedure, was found to be detrimental to the IPG patterns of proteins. The simplified procedure also allowed one to conduct IPG electrophoresis in gel tubes, eliminating lateral zo

ISSN:0173-0835    

DOI:10.1002/elps.1150070604

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

年代:1986

数据来源: WILEY

摘要:

AbstractIn pH 4–10 immobilized pH gradient (IPG) electrophoresis, one observes (a) partial failure of the sample to enter the gel; (b) which halt protein migration; (c) absence of defined transient state band patterns; (d) lateral zone spreading and distorted zone profiles; (d) uneven voltage profiles with a distinct voltage maximum; (f) difficulty in measuring pH profiles with the contact electrode. These problems are remedied through the addition of 0.1 to 1 % carrier ampholytes (CA) to the IPG gel. Problems (a) to (c) disappear in IPG tube gels. IPG in presence of CAs (ICAPG) of pI‐range 3.5–9.5 or 6–8 yields a pH gradient, a protein pattern and a pH gradient stability essentially the same as that in IPG gels of the same pH‐range 4–10. ICAPG is insensitive to ionic strength of the sample up to 0.1 M. Gel swelling and water exudation increase in ICAPG which make ICAPG in tube gels inapplicable when CA concentrations e

ISSN:0173-0835    

DOI:10.1002/elps.1150070605

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

年代:1986

数据来源: WILEY

摘要:

AbstractA sensitive secondary antibody system was developed for detection of small amounts of group specific component (Gc). For the first antibody step after focusig, a comparison between immunofixation on cellulose acetate membranes and capillary blotting on nitrocellulose was made. The sensitivity of the first is much better than that of the latter. The method allowed the detection of about 150 picograms of Gc in 1:12 800 diluted serum samples. Even in six‐year‐old badly soluble bloodstains kept at room temperature, Gc could be phenotyped. The 250 μm thick immobilized pH‐gradient gels for focusing of Gc can be used repeatedly after washing and drying to the initial weight without destroying the pH gr

ISSN:0173-0835    

DOI:10.1002/elps.1150070606

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

年代:1986

数据来源: WILEY

摘要:

AbstractHuman genomic DNA probes labeled by nick translation with biotin 11‐UTP were successfully used to identify single copy restriction fragment length polymorphisms following electrophoresis and Southern blotting onto nylon membranes. Two human probes found on chromosome 21 were used, each of which demonstrated polymorphisms using two separate restriction endonucleases. Compared to isotopic labeling of probes the biotin labels are safer, faster and more economical. The detection method of streptavidin‐biotin‐alkaline phosphatase permitted phenotyping in a matter of hours after hybridization of the probe to the human DNA frag

ISSN:0173-0835    

DOI:10.1002/elps.1150070607

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

年代:1986

数据来源: WILEY

摘要:

AbstractPhenol extracted, alkali‐treated lipopolysaccharide (aLPS) from vaccine strain (S19)Brucella abortuswas demonstrated by two‐dimensional gel electrophoresis to consist of at least ten silver staining, polydisperse analogues having different pIs. When tested on nitrocellulose immunoblots, all ten were antigenically reactive with bovine anti‐B. abortuspolyclonal sera, but only six reacted with anti‐B. abortusO‐antigen murine monoclonal antibody. Analogues focusing at different pIs were concluded to arise from differences in either core or O‐antigen side chain structure or because of covalently bound protein. While not qualitatively different, aLPS from pathogenicB. abortusstrain 2308 had lesser amounts of analogues 1, 2, 5, 6, and 8 than did aLPS from strain 19 (vaccine). The 2‐D gel electrophoresis method was demonstrated to be of value in the analysis of aLPS fromB. abortusand may be useful in the study of lipopolysaccharides from

ISSN:0173-0835    

DOI:10.1002/elps.1150070608

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

年代:1986

数据来源: WILEY

摘要:

AbstractThe FXIIIA polymorphism was studied in a Japanese population using agarose gel isoelectric focusing and agarose gel electrophoresis, combined with immunoblotting onto nitrocellulose membranes. The correspondence of patterns between the two methods was confirmed by two‐dimensional electrophoresis. A rare phenotype, FXIIIA 4–1, was detected in one individual besides the three common phenotypes FXIIIA 1, 2–1, and 2. This rare phenotype was identified with the originally reported FXIIIA 4–1 in Melanesians. Our results indicate the possibility that the FXIIIA*4 allele may be distributed widely in world popu

ISSN:0173-0835    

DOI:10.1002/elps.1150070609

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

年代:1986

数据来源: WILEY

摘要:

AbstractThe electrophoretic behavior of esterase D (E.C. 3.1.1.1.) allozymes 1, 2 and 5 was studied in various buffers and pH ranges, and compared with results obtained on isoelectric focusing. It is shown that the relative mobility of ESD 2 and 5 allozymes is reversed according to the pH of the buffer system. In acidic pH ranges the mobility is: cathode 1<2<5 anode; using the same buffer system in basic pH ranges the following order is observed: 1<5<2. Technical improvements in agarose gel electrophoresis leading to a considerable economical advantage over the previously described methods are presented.

ISSN:0173-0835    

DOI:10.1002/elps.1150070610

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

年代:1986

数据来源: WILEY

ISSN:0173-0835    

DOI:10.1002/elps.1150070611

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

年代:1986

数据来源: WILEY