Biomarker discovery: proteome fractionation and separation in biological samples

2008 ◽  
Vol 33 (1) ◽  
pp. 12-17 ◽  
Author(s):  
Peter Matt ◽  
Zongming Fu ◽  
Qin Fu ◽  
Jennifer E. Van Eyk
Keyword(s):  
Mass Spectrometry ◽  
Biological Samples ◽  
Protein Identification ◽  
Molecular Mechanisms ◽  
Biomarker Discovery ◽  
Clinical Medicine ◽  
Scientific Discipline ◽  
Appropriate Technology ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis

Proteomics, analogous with genomics, is the analysis of the protein complement present in a cell, organ, or organism at any given time. While the genome provides information about the theoretical status of the cellular proteins, the proteome describes the actual content, which ultimately determines the phenotype. The broad application of proteomic technologies in basic science and clinical medicine has the potential to accelerate our understanding of the molecular mechanisms underlying disease and may facilitate the discovery of new drug targets and diagnostic disease markers. Proteomics is a rapidly developing and changing scientific discipline, and the last 5 yr have seen major advances in the underlying techniques as well as expansion into new applications. Core technologies for the separation of proteins and/or peptides are one- and two-dimensional gel electrophoresis and one- and two-dimensional liquid chromatography, and these are coupled almost exclusively with mass spectrometry. Proteomic studies have shown that the most effective analysis of even simple biological samples requires subfractionation and/or enrichment before protein identification by mass spectrometry. Selection of the appropriate technology or combination of technologies to match the biological questions is essential for maximum coverage of the selected subproteome and to ensure both the full interpretation and the downstream utility of the data. In this review, we describe the current technologies for proteome fractionation and separation of biological samples, based on our lab workflow for biomarker discovery and validation.

scholarly journals A Review: Protein Identification by LC-MS: Principles, Instrumentation, and Applications

2020 ◽  
pp. 2448-2466
Author(s):  
Abeer Salh Alhendi
Keyword(s):  
Mass Spectrometry ◽  
Liquid Chromatography ◽  
High Performance ◽  
Protein Identification ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis ◽  
Peptide Separation ◽  
Efficient Separation ◽  
Fundamental Understanding

This review will focus on protein and peptide separation studies of the period 1995 to 2010. Peptide and protein analysis have developed dramatically after applying mass spectrometry (MS) technology and other related techniques, such as two-dimensional liquid chromatography and two-dimensional gel electrophoresis. Mass spectrometry involves measurements of mass-to-charge ratios of the ionized sample. High-performance liquid chromatography (HPLC) is an important technique that is usually applied before MS is conducted due to its efficient separation. Characterization of proteins provides a foundation for the fundamental understanding of biology aspects. In this review, instrumentation, principle, applications, developments, and accuracy of the measurements of mass spectrometry will be reviewed and discussed. In addition, the principles of HPLC technology will be explained, which is necessary before applying MS.

Proteome Analysis of CD4+ T Cells Reveals Differentially Expressed Proteins in Infertile Polycystic Ovary Syndrome Patients

2020 ◽  
Vol 20 ◽  
Author(s):  
Fatemeh Nasri ◽  
Maryam Zare ◽  
Mehrnoosh Doroudchi ◽  
Behrouz Gharesi-Fard
Keyword(s):  
Mass Spectrometry ◽  
T Cells ◽  
T Lymphocytes ◽  
Gel Electrophoresis ◽  
Cd4 T Cells ◽  
Polycystic Ovary ◽  
Proteome Analysis ◽  
Two Dimensional ◽  
Ovary Syndrome ◽  
Two Dimensional Gel Electrophoresis

Background: Polycystic ovary syndrome (PCOS) is the most frequent endocrine disorder affecting 6–7% of premenopausal women. Recent studies revealed that the immune system especially CD4+ T helper cells are important in the context PCOS. Proteome analysis of CD4+ T lymphocytes can provide valuable information regarding the biology of these cells in the context of PCOS. Objective: To investigate immune dysregulation in CD4+ T lymphocytes at the protein level in the context of PCOS using two-dimensional gel electrophoresis (2DE) and mass spectrometry (MS). Methods: In the present study, we applied two-dimensional gel electrophoresis / mass spectrometry to identify proteins differentially expressed by peripheral blood CD4+ T cells in ten PCOS women compared with ten healthy women. Western blot technique was used to confirm the identified proteins. Results: Despite the overall proteome similarities, there were significant differences in the expression of seven spots between two groups (P <0.05). Three proteins, namely phosphatidylethanolamine-binding protein 1, proteasome activator complex subunit 1 and triosephosphate isomerase 1 were successfully identified by Mass technique and confirmed by western blot. All characterized proteins were over-expressed in CD4+ T cells from patients compared to CD4+ T cells from controls (P <0.05). In-silico analysis suggested that the over-expressed proteins interact with other proteins involved in cellular metabolism especially glycolysis and ferroptosis pathway. Conclusion: These findings suggest that metabolic adjustments in CD4+ T lymphocytes, which is in favor of increased glycolysis and Th2 differentiation are important in the context of PCOS.

Two-dimensional electrophoresis of human placental mitochondria and protein identification by mass spectrometry: Toward a human mitochondrial proteome

1998 ◽  
Vol 19 (6) ◽  
pp. 1006-1014 ◽  
Author(s):  
Thierry Rabilloud ◽  
Sylvie Kieffer ◽  
Vincent Procaccio ◽  
Mathilde Louwagie ◽  
Paul L. Courchesne ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Protein Identification ◽  
Two Dimensional ◽  
Mitochondrial Proteome ◽  
Two Dimensional Electrophoresis ◽  
Dimensional Electrophoresis

scholarly journals Proteomic analysis of the pulvinus, a heliotropic tissue, in Glycine max

2014 ◽  
Vol 5 (1) ◽  
Author(s):  
Hakme Lee ◽  
Wesley M. Garrett ◽  
Joseph Sullivan ◽  
Irwin Forseth ◽  
Savithiry S. Natarajan
Keyword(s):  
Mass Spectrometry ◽  
Gene Ontology ◽  
Gel Electrophoresis ◽  
Proton Transport ◽  
Enrichment Analysis ◽  
The Sun ◽  
Leaf Movement ◽  
Two Dimensional ◽  
Two Dimensional Gel Electrophoresis ◽  
Leguminous Plants

Certain plant species respond to light, dark, and other environmental factors by leaf movement. Leguminous plants both track and avoid the sun through turgor changes of the pulvinus tissue at the base of leaves. Mechanisms leading to pulvinar turgor flux, particularly knowledge of the proteins involved, are not well-known. In this study we used two-dimensional gel electrophoresis and liquid chromatography-tandom mass spectrometry to separate and identify the proteins located in the soybean pulvinus. A total of 183 spots were separated and 195 proteins from 165 spots were identified and functionally analyzed using single enrichment analysis for gene ontology terms. The most significant terms were related to proton transport. Comparison with guard cell proteomes revealed similar significant processes but a greater number of pulvinus proteins are required for comparable analysis. To our knowledge, this is a novel report on the analysis of proteins found in soybean pulvinus. These findings provide a better understanding of the proteins required for turgor change in the pulvinus.

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