scholarly journals Two-dimensional liquid chromatography protein expression mapping for differential proteomic analysis of normal and O157:H7 Escherichia coli

BioTechniques ◽  
2003 ◽  
Vol 35 (6) ◽  
pp. 1202-1212 ◽  
Author(s):  
Suping Zheng ◽  
Kimberly A. Schneider ◽  
Timothy J. Barder ◽  
David M. Lubman
Keyword(s):  
Escherichia Coli ◽  
Liquid Chromatography ◽  
Protein Expression ◽  
Proteomic Analysis ◽  
Two Dimensional

Proteomic analysis reveals protein expression differences in Escherichia coli strains associated with persistent versus transient mastitis

2014 ◽  
Vol 108 ◽  
pp. 373-381 ◽  
Author(s):  
John D. Lippolis ◽  
Brian W. Brunelle ◽  
Timothy A. Reinhardt ◽  
Randy E. Sacco ◽  
Brian J. Nonnecke ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Protein Expression ◽  
Proteomic Analysis

scholarly journals Rapid Proteome Analysis of Bronchoalveolar Lavage Samples of Lifelong Smokers and Never-Smokers by Micro-Scale Liquid Chromatography and Mass Spectrometry

2006 ◽  
Vol 52 (4) ◽  
pp. 671-679 ◽  
Author(s):  
Amelie Plymoth ◽  
Ziping Yang ◽  
Claes-Göran Löfdahl ◽  
Ann Ekberg-Jansson ◽  
Magnus Dahlbäck ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Liquid Chromatography ◽  
Protein Expression ◽  
Bronchoalveolar Lavage ◽  
Proteomic Analysis ◽  
Ion Trap ◽  
Wide Spectrum ◽  
Proteome Analysis ◽  
Chronic Obstructive ◽  
Never Smokers

Abstract Background: The aim of this study was to determine whether relative qualitative and quantitative differences in protein expression could be related to smoke exposure or smoke-induced airway inflammation. We therefore explored and characterized the protein components found in bronchoalveolar lavage (BAL) fluid sampled from either lifelong smokers or never-smokers. Methods: BAL fluid samples obtained by bronchoscopy from 60-year-old healthy never-smokers (n = 18) and asymptomatic smokers (n = 30) were analyzed in either pooled or individual form. Initial global proteomic analysis used shotgun digestion approaches on unfractionated BAL fluid samples (after minimal sample preparation) and separation of peptides by gradient (90-min) liquid chromatography (LC) coupled with on-line linear ion trap quadropole mass spectrometry (LTQ MS) for identification and analysis. Results: LTQ MS identified 481 high- to low-abundance proteins. Relative differences in patterns of BAL fluid proteins in smokers compared with never-smokers were observed in pooled and individual samples as well as by 2-dimensional gel analysis. Gene ontology categorization of all annotated proteins showed a wide spectrum of molecular functions and biological processes. Conclusions: The described method provides comprehensive qualitative proteomic analysis of BAL fluid protein expression from never-smokers and from smokers at risk of developing chronic obstructive pulmonary disease. Many of the proteins identified had not been detected in previous studies of BAL fluid; thus, the use of LC-tandem MS with LTQ may provide new information regarding potentially important patterns of protein expression associated with lifelong smoking.

62 COMPARATIVE PROTEOMIC ANALYSIS OF LIVER MITOCHONDRIA DERIVED FROM DECEASED NEWBORN CLONED CALVES AND ADULT CLONES BY TWO-DIMENSIONAL DIFFERENTIAL GEL ELECTROPHORESIS

2011 ◽  
Vol 23 (1) ◽  
pp. 137
Author(s):  
K. Takeda ◽  
M. Tasai ◽  
S. Akagi ◽  
S. Watanabe ◽  
M. Oe ◽  
...  
Keyword(s):  
Protein Expression ◽  
Somatic Cell ◽  
Proteomic Analysis ◽  
Gel Electrophoresis ◽  
Mitochondrial Protein ◽  
Two Dimensional ◽  
Cell Nuclei ◽  
Comparative Proteomic Analysis ◽  
Mitochondrial Fractions

Aberrant reprogramming of donor somatic cell nuclei may result in many severe problems in animal cloning. The inability to establish functional interactions between donor nucleus and recipient mitochondria is also likely responsible for developmental deficiency. However, an understanding of the expressed proteins in cattle is lacking. In the present study, alterations in mitochondrial protein levels between somatic cell nuclear transferred (SCNT) and control animals (mostly produced by AI) were investigated. Nuclear transfer was performed using donor cells prepared from cumulus cells (B1), ear skin, or skeletal muscle from adult Japanese Black cattle, and enucleated in vitro matured oocytes (Holstein or Japanese Black) as previously reported (Akagi et al. 2003). Liver samples were collected from postmortem SCNT calves (CB1-3; 0, 1, and 9 days postnatally) and adult SCNT cattle (CA1-4; 6, 6, 6, and 5 years of age) produced from the same cell line (B1) and preserved at –80°C. Mitochondrial fractions were prepared from the frozen–thawed liver samples by mechanical homogenization and differential centrifugation, and subjected to two-dimensional difference in gel electrophoresis (2D-DIGE) using CyDye™ dyes (Cy2, Cy3, Cy5; GE Healthcare) for specific labelling. Protein expression changes were confirmed by ImageMaster 2D Platinum software with a volume ratio greater than 2.0 (Student’s t-test; P < 0.05). The expression of 5 proteins were up-regulated in SCNT calves compared to control calves (n = 6; Day 250 fetus, 0, 4, 8, 8, and 8 days after birth; P < 0.05). Expressed protein patterning compared to control groups was different among SCNT calves. The protein spots of CB-1 showed great differences compared with other SCNT calves; 13 spots were up-regulated, and 18 spots were down-regulated. In adult SCNT cattle, the concentrations of 3 proteins were higher when compared to control cattle (n = 4; 2, 2, 6, and 8 years of age; P < 0.05). Protein expression was different among individual SCNT animals even if they were produced from the same donor cell source. For example, 9 spots were up-regulated and 7 spots were down-regulated in CA-1. In contrast, no differences were detected in 2 of the SCNT cattle (CA-3 and 4; P < 0.05). Novel proteins were not identified in any of the SCNT cattle or calves. In conclusion, alteration of mitochondrial protein expression levels were observed in non-viable neonatal SCNT calves and varied among SCNT individuals; suggesting that mitochondrial related gene expression may be implicated in early losses. Comparative proteomic analysis represents an important tool for further studies on SCNT animals. We thank Dr. C. A. Pinkert (Auburn Univ.) and Dr. Somfai (NARO) for their assistance. This work was supported by a grant from the NARO, Japan.

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