Strategies for the Isolation of Peptides from Low-Abundance Proteins for Internal Sequence Analysis33Work was supported in part by the Genetics Network of Centres of Excellence (Canada) and by the Biomedical Research Centre. R.A. was the recipient of an MRC of Canada Scholarship. D.H. was the recipient of a fellowship from the Swiss National Science FoundationAbbreviations: BNPS-skatole: 2-(2’-nitrophenylsulfenyl)-3-methyl-3’-bromoindolenine; BSA: bovine serum albumin; CAPS: 3-[cyclohexylamino]-1-propanesulfonic acid; CD: Cationic Durapore; CNBr: cyanogen bromide; GuHCl: Guanidine hydrochloride; IPG-IEF: immobilized pH gradient isoelectric focusing; NC: nitrocellulose; PVDF: polyvinylidene fluoride; PVP: polyvinylpyrrolidone; RP-HPLC: reverse phase-high performance liquid chromatography; SDS-PAGE: sodium dodecyl sulfate polsyacrylamide gel electrophoresis; 2DE: two-dimensional gel electrophoresis; TFA: trifluoroacetic acid.

1992 ◽  
pp. 87-96 ◽  
Author(s):  
Ruedi Aebersold ◽  
Scott D. Patterson ◽  
Daniel Hess
Keyword(s):  
Gel Electrophoresis ◽  
Polyvinylidene Fluoride ◽  
High Performance ◽  
Guanidine Hydrochloride ◽  
Sodium Dodecyl ◽  
Research Centre ◽  
Two Dimensional Gel Electrophoresis ◽  
Internal Sequence ◽  
Sds Page ◽  
Centres Of Excellence

Study on Degradation of Radix Isatidis Polypeptide in Artificial Gastrointestinal Environment

2014 ◽  
Vol 989-994 ◽  
pp. 1020-1024
Author(s):  
Nan Nan ◽  
Xi Jing Liu
Keyword(s):  
Gel Electrophoresis ◽  
Free Amino ◽  
High Performance ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Radix Isatidis ◽  
Sds Page ◽  
Electrophoresis Method ◽  
Amino Acid Levels ◽  
Different Molecular Weight

Radix Isatidis is a traditional Chinese medicine for treatment of influenza and inflammation in China. In this paper, in order to study the degradation situation of Radix Isatidis polypeptide in artificial gastrointestinal environment, the SDS-PAGE (Sodium dodecyl sulfate-polyacrylamide gel electrophoresis) method was used to detect the degradation of Radix Isatidis polypeptide in artificial intestinal juice and gastric juice, and it showed that Radix Isatidis peptides could be degradated to different degrees. HPLC (High Performance Liquid Chromatography) was used to determine the change of peptides degradation, and it indicated that free amino acid levels did not change significantly. The result after degradation was also detected by BCA method, and it showed that there were still a large number of polypeptides in the liquid. From this experiment we can come to this conclusion that Radix Isatidis polypeptides in artificial gastrointestinal juice mostly degraded into a series of different molecular weight peptides.

Sugar profiling proves that human serum erythropoietin differs from recombinant human erythropoietin

Blood ◽  
2001 ◽  
Vol 98 (13) ◽  
pp. 3626-3634 ◽  
Author(s):  
Venke Skibeli ◽  
Gro Nissen-Lie ◽  
Peter Torjesen
Keyword(s):  
Human Serum ◽  
Gel Electrophoresis ◽  
High Performance ◽  
Magnetic Beads ◽  
Direct Method ◽  
Polyacrylamide Gel Electrophoresis ◽  
Broad Band ◽  
Sodium Dodecyl ◽  
Apparent Molecular Weight ◽  
Two Dimensional Gel Electrophoresis

Abstract Erythropoietin (EPO) from sera obtained from anemic patients was successfully isolated using magnetic beads coated with a human EPO (hEPO)–specific antibody. Human serum EPO emerged as a broad band after sodium dodecyl sulfate–polyacrylamide gel electrophoresis, with an apparent molecular weight slightly smaller than that of recombinant hEPO (rhEPO). The bandwidth corresponded with microheterogeneity because of extensive glycosylation. Two-dimensional gel electrophoresis revealing several different glycoforms confirmed the heterogeneity of circulating hEPO. The immobilized anti-hEPO antibody was capable of binding a representative selection of rhEPO glycoforms. This was shown by comparing normal-phase high-performance liquid chromatography profiles of oligosaccharides released from rhEPO with oligosaccharides released from rhEPO after isolation with hEPO-specific magnetic beads. Charge analysis demonstrated that human serum EPO contained only mono-, di-, and tri-acidic oligosaccharides and lacked the tetra-acidic structures present in the glycans from rhEPO. Determination of charge state after treatment of human serum EPO with Arthrobacter ureafaciens sialidase showed that the acidity of the oligosaccharide structures was caused by sialic acids. The sugar profiles of human serum EPO, describing both neutral and charged sugar, appeared significantly different from the profiles of rhEPO. The detection of glycan structural discrepancies between human serum EPO and rhEPO by sugar profiling may be significant for diagnosing pathologic conditions, maintaining pharmaceutical quality control, and establishing a direct method to detect the misuse of rhEPO in sports.

Fractionation of nucleolar proteins by two-dimensional gel electrophoresis

1976 ◽  
Vol 54 (1) ◽  
pp. 9-14 ◽  
Author(s):  
G. Jackowski ◽  
D. Suria ◽  
C. C. Liew
Keyword(s):  
Sodium Dodecyl Sulfate ◽  
Gel Electrophoresis ◽  
Guanidine Hydrochloride ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis ◽  
Nucleolar Proteins ◽  
Dodecyl Sulfate ◽  
Ph Range

Isolation of nucleolar proteins was obtained by dissociation in the presence of urea – guanidine hydrochloride, followed by high-speed centrifugation to remove nucleic acids. At least 31 fractions of nucleolar proteins were detected by isoelectrofocusing gel electrophoresis in the pH range 3.5–10. Following two-dimensional gel electrophoresis on sodium dodecyl sulfate – polyacrylamide slab gels, more than 100 components of nucleolar proteins were identified. Two-thirds of nucleolar proteins were located in the pH range 5–8 following isoelectrofocusing. The molecular weights of these classes of proteins were shown to be mostly 30 000 – 70 000 by sodium dodecyl sulfate – polyacrylamide gel electrophoresis.

scholarly journals Demonstration of structural polymorphism among HLA-DR light chains by two-dimensional gel electrophoresis.

1980 ◽  
Vol 151 (1) ◽  
pp. 144-165 ◽  
Author(s):  
D A Shackelford ◽  
J L Strominger
Keyword(s):  
Cell Lines ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Light Chains ◽  
Two Dimensional ◽  
Structural Polymorphism ◽  
Two Dimensional Gel Electrophoresis ◽  
Hla Dr ◽  
Sds Page

Human HLA-DR antigens were immunoprecipitated from Nonidet P-40 extracts of [35S]methionine-labeled B lymphoblastoid cell lines and compared by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and isoelectric focusing (IEF). Two-dimensional (2-D) gel analyses, combining SDS-PAGE in the first dimension and IEF in the second dimension, revealed that the heavy (alpha) and light (beta) chains of each DRw specificity displays microheterogeneity of charge. However, the pattern of the heavy chain did not vary among different DRw specificities. In contrast, the light chains of different DRw types varied both in apparent size and charge distribution. Removal of sialic acids with neuraminidase or inhibition of glycosylation with tunicamycin reduced the microheterogeneity of both DR subunits. However, the heavy and light chains each still focused as two major bands, suggesting that other post-translational modifications contribute to the microheterogeneity or that there are two nonallelic DR-like molecules. After treatment with either neuraminidase or tunicamycin, the DR light chains, but not the heavy chains, were still structurally polymorphic. The DR light chains of serologically cross-reactive specificities displayed similar 2-D gel patterns suggesting that the structural polymorphism of the DR light chains is the basis for the serologically detected polymorphism of the HLA-DR antigens. Two additional polypeptides were observed in immunoprecipitates of DR antigens. These proteins, designated M1 and M2, both had a basic isoelectric point and were invariant among different cell lines. The protein M1 may be intracellular because it can not be immunoprecipitated from the cell surface.

Two-dimensional gel electrophoresis method for investigation of human plasma proteins: detection of subtle changes during filtration leukapheresis.

1982 ◽  
Vol 28 (4) ◽  
pp. 962-968 ◽  
Author(s):  
K Lonberg-Holm ◽  
E A Bagley ◽  
J Nusbacher ◽  
J M Heal
Keyword(s):  
Human Plasma ◽  
Gel Electrophoresis ◽  
Plasma Proteins ◽  
Acute Phase Proteins ◽  
Venous Blood ◽  
Sodium Dodecyl ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis ◽  
Native Proteins ◽  
Sds Page

Abstract Special problems are associated with analysis of human plasma proteins by standard "high-resolution" two-dimensional gel electrophoresis methods in which isoelectric focusing is followed by electrophoresis in sodium dodecyl sulfate/polyacrylamide gels (SDS-PAGE). Individual plasma proteins are often separated into overlapping groups of multiple spots, and identification of individual spots is further confounded by genetic variation. Analytical recovery of components of high molecular mass is also low or variable. These problems may be reduced or overcome by use of a "low-resolution" method consisting of electrophoresis of native proteins at pH 8.6 in an agarose gel followed by SDS-PAGE without added reducing agent. About 60 proteins can be resolved, most as single spots. About 25 of these proteins have been "mapped," and tentatively identified. We have examined 119 plasma samples taken from six donors who were undergoing filtration leukapheresis and 10 donors who were undergoing centrifugation leukapheresis or plateletpheresis. In all cases, passage of blood through a nylon filter induced a significant increase in a doublet of spots tentatively identified as complement component C3c. This was detected in the effluent from the filter throughout the first 30 min of filtration, and to a lesser extent in the venous blood. These spots were not induced by the centrifugation procedures. One filtration donor also showed increased acute-phase proteins 24 h after the procedure.

Heterozygous Abnormal Fibrinogen Osaka III with the Replacement of γ Arginine-275 by Histidine Has an Apparently Higher Molecular Weight γ-Chain Variant

1992 ◽  
Vol 68 (05) ◽  
pp. 534-538 ◽  
Author(s):  
Nobuhiko Yoshida ◽  
Shingi Imaoka ◽  
Hajime Hirata ◽  
Michio Matsuda ◽  
Shinji Asakura
Keyword(s):  
Molecular Weight ◽  
Sodium Dodecyl Sulfate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Tetraacetic Acid ◽  
Fibrin Monomer ◽  
Sds Page ◽  
Thrombotic Tendency ◽  
Chain Variant

SummaryCongenitally abnormal fibrinogen Osaka III with the replacement of γ Arg-275 by His was found in a 38-year-old female with no bleeding or thrombotic tendency. Release of fibrinopeptide(s) by thrombin or reptilase was normal, but her thrombin or reptilase time in the absence of calcium was markedly prolonged and the polymerization of preformed fibrin monomer which was prepared by the treatment of fibrinogen with thrombin or reptilase was also markedly defective. Propositus' fibrinogen had normal crosslinking abilities of α- and γ-chains. Analysis of fibrinogen chains on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in the system of Laemmli only revealed the presence of abnormal γ-chain with an apparently higher molecular weight, the presence of which was more clearly detected with SDS-PAGE of fibrin monomer obtained by thrombin treatment. Purified fragment D1 of fibrinogen Osaka III also seemed to contain an apparently higher molecular weight fragment D1 γ remnant on Laemmli gels, which was digested faster than the normal control by plasmin in the presence of [ethy-lenebis(oxyethylenenitrilo)]tetraacetic acid (EGTA).

Identifikasi Daging Tikus Pada Produk Baso Dengan Metode Sodium Dodecyl Sulphate Polyacrylamide Gel Electrophoresis (SDS-PAGE)

2018 ◽  
Vol 26 (2) ◽  
pp. 058
Author(s):  
Anna P. Roswiem ◽  
Triayu Septiani
Keyword(s):  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Polyacrylamide Gel ◽  
Sds Page

<em>Bahan<strong> </strong>baku untuk membuat baso adalah daging hewan, pada umumnya dari daging sapi, ayam, ikan dan babi. Di beberapa daerah di Indonesia terjadi kasus baso tikus. Tujuan penelitian ini adalah menguji ada tidaknya kandungan daging tikus pada produk baso yang dijual di pasar Cempaka Putih-Kecamatan Kramat Jakarta Pusat dan di pedagang baso atau mie baso di sekitar kampus Universitas YARSI Jakarta. Daging adalah protein salah satu metode untuk mengidentifikasi protein adalah metode Sodium Dodecyl Sulphate Polyacrylamide Gel Electrophoresis (SDS-PAGE).<strong> </strong>Hasil penelitian menunjukkan bahwa dari 6 sampel baso terindikasi ada 2 sampel baso dengan nomor 1 dan 5 yang dibuat dari campuran daging sapi dan tikus; ada 1 sampel baso dengan nomor 6 yang terbuat dari daging tikus; dan 2 sampel baso dengan nomor 2 dan 3 yang terbuat dari campuran sapi  dan babi, dan hanya 1 sampel baso dengan nomor sampel 4 yang benar-benar terbuat dari daging sapi.</em>

Effect of detergents and chemicals on purified vaccinia virus: analysis by SDS polyacrylamide gel electrophoresis and electron microscopy

1977 ◽  
Vol 23 (3) ◽  
pp. 240-252 ◽  
Author(s):  
J. Boisvert ◽  
T. Yamamoto
Keyword(s):  
Electron Microscopy ◽  
Vaccinia Virus ◽  
Gel Electrophoresis ◽  
Alkali Treatment ◽  
Guanidine Hydrochloride ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Ammonium Bromide ◽  
Molecular Weights ◽  
Viral Particles

Vaccinia virus particles were dissociated into their constituent polypeptides and analysed by sodium dodecyl sulfate (SDS) gel electrophoresis. Thirty-three distinct polypeptide bands were identified and their molecular weights ranged between 11 000 and 150 000 daltons.Specific staining of gels containing polypeptides of dissociated virions revealed the presence of eight glycopeptides. No lipopeptides were detected.Analysis of chemical extracts (urea, guanidine hydrochloride, and alkali treatment) of the virus by SDS gel electrophoresis indicated that a total of 10 to 14 different polypeptides ranging in molecular weights from 11 000 to 70 000 daltons were solubilized.Analysis of detergent extracts and of the remains of extracted viral particles has shown that the detergent Nonidet P-40 (NP-40) solubilized a total of 11 polypeptides of which 6 were glycopeptides. The other detergents sodium deoxycholate (SDC) and cetyl trimethyl ammonium bromide (CTAB) were not as selective, both solubilizing more than 25 of the polypeptides composing the virus. Gel electrophoresis results also indicated that most of the small molecular weight (11 000–70 000 daltons) polypeptides were readily solubilized by NP-40, SDC, and CTAB, while those with molecular weights of 70 000 daltons and higher were not well solubilized.The effects of detergents were also analysed by electron microscopy. Evidence was obtained for subpopulations of viral particles having different susceptibility to detergent extraction.

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