Two-dimensional gel electrophoresis of cerebrospinal fluid proteins.

1980 ◽  
Vol 26 (9) ◽  
pp. 1317-1322 ◽  
Author(s):  
D Goldman ◽  
C R Merril ◽  
M H Ebert
Keyword(s):  
Cerebrospinal Fluid ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Spinal Fluid ◽  
Two Dimensional ◽  
Silver Stain ◽  
Two Dimensional Gel Electrophoresis ◽  
Cerebrospinal Fluid Proteins ◽  
High Resolution Analysis

Abstract Two-dimensional electrophoresis, with isoelectric focusing in the first dimension and sodium dodecyl sulfate/polyacrylamide gel electrophoresis in the second, has been adapted for the high-resolution analysis of cerebrospinal fluid proteins. Proteins were detected with a new, highly sensitive silver stain that made visible more than 300 polypeptides from 60 microL of spinal fluid, in highly reproducible patterns. We have mapped these patterns, noting difference between the proteins observed in spinal fluid and plasma, and have prepared a partial map of cerebrospinal fluid proteins.

Reproducibility and quality assurance of two-dimensional gel electrophoresis of serum specimens.

1982 ◽  
Vol 28 (4) ◽  
pp. 908-914 ◽  
Author(s):  
R P Tracy ◽  
R M Currie ◽  
D S Young
Keyword(s):  
Quality Assurance ◽  
Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Two Dimensional ◽  
Protein Loss ◽  
Serum Samples ◽  
Silver Stain ◽  
Two Dimensional Gel Electrophoresis ◽  
Report Quality ◽  
Wide Range

Abstract Currently we are using two different ISO-DALT two-dimensional gel electrophoresis systems, designated MC-Iso 1 and MC-Iso 2, for the analysis of serum and plasma samples. Here we report quality-assurance data for both of these systems. CV values for the slopes of the pH gradient (ISO dimension) are 5.6% of less; CV values for the slopes of the molecular-mass curves (log Mr vs relative mobility in the DALT dimension) are 3.4% or less. We examined the various steps of the analysis in detail for reproducibility and protein loss, using radiolabeled albumin, alpha 2-macroglobulin, and beta 2-microglobulin. Generally, in the first dimension, less protein enters the MC-Iso 2 gels (our routine system in which silver stain is used) than enters the MC-Iso 1 gels (our wide-range system for myeloma serum samples, in which the gel is stained with Coomassie Blue), on the average, 87% as much. The CV at this stage for both systems is 5--8%. During equilibration, considerable amounts of protein are lost (approximately 30% in 10 min) from the ISO gel, and the reproducibility is also decreased. Resolution in the DALT dimension has, in most cases, little or no effect on either recovery or reproducibility. Overall, for most proteins expected to appear in an ISO gel of a given pH range, approximately 50--60% of the starting material may be expected to reside in the sodium dodecyl sulfate slab gel, under our conditions. The two most important variables affecting recovery are the concentration of the NaOH (used as catholyte) and the pH of the starting sample. The overall CV for the process is between 8 and 12%.

A New Method for Proteome Screening with Two-Dimensional Urea-Sds Polyacrylamide Gel Electrophoresis

2014 ◽  
Vol 1 (1) ◽  
Author(s):  
Areeba Ahmad ◽  
Riaz Ahmad
Keyword(s):  
Gel Electrophoresis ◽  
Protein Separation ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Cost Effective ◽  
Polyacrylamide Gel ◽  
Present System ◽  
Liver Protein ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis

AbstractTwo-dimensional gel electrophoresis (2DE) separating proteins on the basis of their pI and molecular mass remain the best available technique for protein separation and characterization to date. But due to several limitations, including streak formation in IEF gels, partial solubility of proteins, expensive running conditions and relatively longer time taken, a simple urea-SDS-2D polyacrylamide gel electrophoresis (US2DE) is described here. The system is reasonably sensitive, cost effective with good reproducibility. The method described in this paper employs a chaotropic agent, urea, in the first dimension and sodium dodecyl sulphate (SDS), like conventional system, in the second dimension with an addition of polyacrylamide to screen the liver proteome of healthy and chemically induced fibrotic rats. The system separates the protein on the basis of chargeto- mass ratio and clearly demonstrates differential expression in the liver protein repertoire of healthy and fibrotic rats. Moreover, the present system, like other 2D electrophoretic procedures revealed at least 22 novel spots in the investigated tissues. The technique may be utilized for comprehensive proteome screening of any biological sample and would provide an overview to narrow down the candidate proteins or biomarkers.

scholarly journals A novel two-dimensional polyacrylamide-gel pattern, which may be due to allelic genes, of phenylalanine hydroxylase in monkeys

1985 ◽  
Vol 231 (1) ◽  
pp. 197-199 ◽  
Author(s):  
S C Smith ◽  
W McAdam ◽  
R G H Cotton ◽  
J F B Mercer
Keyword(s):  
Isoelectric Focusing ◽  
Gel Electrophoresis ◽  
Phenylalanine Hydroxylase ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Polyacrylamide Gel ◽  
The Novel ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis ◽  
Monkey Liver

Two-dimensional gel electrophoresis of immunopurified monkey liver phenylalanine hydroxylase showed a novel form of the enzyme, in 4 out of 24 monkeys, in which each polypeptide spot was split into a doublet with the same charge but slightly different mobility in the sodium dodecyl sulphate/polyacrylamide-gel-electrophoresis (as opposed to the isoelectric-focusing) dimension. Phenylalanine hydroxylase formed by translation of RNA from a liver containing the novel form showed the doublet pattern, suggesting that it is due to differences in mRNA. By analogy with the rat, this mRNA difference could be due to allelic genes.

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.

scholarly journals Evaluation of proteome profiles of Salix spp. pollen and relationship between glucose oxidase activity and pollen content in willow honey

2019 ◽  
Vol 25 (1) ◽  
Author(s):  
Violeta Leonora Čeksterytė ◽  
V Borutinskaitė ◽  
D Matuzevičius ◽  
G Treigytė ◽  
D Navakauskas ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Glucose Oxidase ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis ◽  
Pollen Content ◽  
2D Gel ◽  
Salix Spp

Proteins from hand- and bee- collected pollen were isolated and separated in sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and two-dimensional gel electrophoresis (2DE) system as well by direct gel-free mass spectrometry (MS) analysis. Total thirty six plant proteins with known functions and three uncharacterized were identified using mass spectrometry (MS) techniques from fourteen protein spots separated in 2D gel electrophoresis. Our study revealed two forms of profilin proteins with their slight different mass 14.19 kDa and 14.09 kDa and beta actin fragment was 33.25 kDa both in hand and bee collected pollen as well proteins of thioredoxin family with molecular weight - 47.29 kDa.  The antioxidant enzyme glucose oxidase (GOX) activity was investigated in twelve samples of monofloral willow honey, having Salix spp. pollen from 44.7 % to 80.2 %. Data of paired regression analysis between Salix spp. pollen content and GOX activity show moderate and negative correlation coefficient (r= - 0.56). Keywords: willow pollen; honey; protein expression; separation; two-dimensional electrophoresis; mass spectrometry

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