ISSN:0173-0835    

DOI:10.1002/elps.1150020202

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

年代:1981

数据来源: WILEY

摘要:

AbstractA new fluorescent staining technique using o‐phthaldialdehyde(OPA) has been developed for the rapid post‐electrophoretic location for separated zones of proteins and glycoproteins. The method has also been applied to the detection of immunoprecipitates and, in the form of a spray reagent, to the detection of proteins separated on thin‐layer gel filtration plates. Major protein zones can be visualized within 1–2 min with maximum sensitivity being achieved within 15 min. In polyacrylamide and agarose gels, the sensitivity is of the same order as staining with Coomassie Blue G‐250. Reagents are very inexpensive and, because the reaction is performed at neutral or weekly alkaline pH in aqueous buffers, retention of enzymatic or other biological activity should be optimal. The method is thus particularly suitable for use in preparative applications and when it is desirable to combine general protein detection with specific location of zone of enzyme

ISSN:0173-0835    

DOI:10.1002/elps.1150020203

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

年代:1981

数据来源: WILEY

摘要:

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

ISSN:0173-0835    

DOI:10.1002/elps.1150020204

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

年代:1981

数据来源: WILEY

摘要:

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

ISSN:0173-0835    

DOI:10.1002/elps.1150020205

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

年代:1981

数据来源: WILEY

摘要:

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 orStaphylococcusV8 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.StaphylococcusV8 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

ISSN:0173-0835    

DOI:10.1002/elps.1150020206

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

年代:1981

数据来源: WILEY

摘要:

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 sour

ISSN:0173-0835    

DOI:10.1002/elps.1150020207

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

年代:1981

数据来源: WILEY

摘要:

AbstractA two‐dimensional crossed immunoelectrophoretic method for the determination of the specificity of antibodies raised against rat serum lipoproteins and apolipoproteins (Apo) is described. The method may also be used for the detection of as yet undesignated apoproteins, associated with the serum lipoprotein fractions, and exploits the superior resolving capacity of 7m urea 10% polyacrylamide gel electrophoresis for the first dimension separation and minimizes physical separation of polyacrylamide and agarose in the second dimension electrophoresis by use of low electroendosmotic flow agarose. The use of polyacrylamide instead of cellulose acetate in the first dimension permits the analysis of larger amounts of antigen protein, crossed imunoelectrophoresis of Apo E which binds to cellulose acetate, and the crossed immunoelectrophoresis of antigens previously focused to their isoelectric points in polyacrylamide gels containing ampholyte

ISSN:0173-0835    

DOI:10.1002/elps.1150020208

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

年代:1981

数据来源: WILEY

摘要:

AbstractCapillary isotachophoresis has been applied as an analytical monitor during the synthesis of the C‐terminal pentapeptide, Gln‐His‐Phe‐Ala‐Asn, of bombinin. The samples were separated as cations since the C‐terminal carboxyl group was modified to an uncharged amide. Potassium and beeta‐alanine were employed as leading and terminating ions and acetate was the common counter‐ion. The analyses demonstrated that the stages of synthesis up to the tripeptide, Phe‐Ala‐Asn, gave quite pure products, whereas a considerable amount of impurity was detectable after the introduction of His into the molecule. From the peptide chemist's point of view, this amino acid presents special problems. However, the impurities found in the isotachopherograms were not detectable by conventional methods such as thin‐lyer chromatography. The rapid and sensitive method of isotachophoresis should prove useful to the peptide chemist for assessment of his products, and for redesign or modification of non‐optim

ISSN:0173-0835    

DOI:10.1002/elps.1150020209

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

年代:1981

数据来源: WILEY

摘要:

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 tota

ISSN:0173-0835    

DOI:10.1002/elps.1150020210

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

年代:1981

数据来源: WILEY

ISSN:0173-0835    

DOI:10.1002/elps.1150020211

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

年代:1981

数据来源: WILEY