Nanoscale proteomics

Shen, Yufeng; Tolić, Nikola; Masselon, Christophe; Paša-Tolić, L.; Camp, David G.; Lipton, Mary; Anderson, Gordon A.; Smith, Richard D.

doi:10.1007/s00216-003-2329-8

Cited by 53 publications

(44 citation statements)

References 24 publications

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“…One particularly efficient method for proteome analysis is the so-called accurate mass and time (AMT) tag approach where high performance liquid chromatography (HPLC) and high resolution Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS or FTMS) work in synergy to achieve a broad proteome coverage even for small sample amounts (20,21). A recent report demonstrated the significant advantage of the AMT tag method over more conventional proteomics technologies in enabling significantly broader proteome coverage using the minute protein quantities available from tissue microdissection (22).…”

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confidence: 99%

Identification of Cellular Targets in Human Intrahepatic Cholangiocarcinoma Using Laser Microdissection and Accurate Mass and Time Tag Proteomics

Santos

Court

Thiers

et al. 2010

Molecular & Cellular Proteomics

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Obtaining accurate protein profiles from homogeneous cell populations in heterogeneous tissues can enhance the capability to discover protein biomarkers. In this context, methodologies to access specific cellular populations and analyze their proteome with exquisite sensitivity have to be selected. We report here the results of an investigation using a combination of laser microdissection and accurate mass and time tag proteomics. The study was aimed at the precise determination of proteome alterations in intrahepatic cholangiocarcinoma ICC, a markedly heterogeneous tumor. This cancer, which is difficult to diagnose and carries a very poor prognosis, has shown an unexplained increase in incidence over the last few years. Among a pool of 574 identified proteins, we were able to report on altered abundance patterns affecting 39 proteins conforming to a variety of potential tumorigenic pathways. The reliability of the proteomics results was confirmed by Western blot and immunohistochemistry on matched samples. Most of the proteins displaying perturbed abundances had not yet been described in the setting of ICC. These include proteins involved in cell mobility and actin cytoskeleton remodeling, which may participate in the epithelial to mesenchymal transition, a process invoked in migration and invasion of cancer cells. The biological relevance of these findings was explored using a tissue microarray. An increased abundance of vimentin was thus detected in 70% of ICC and none of the controls. These results suggest that vimentin could play a role in the aggressiveness of ICC and provide a basis for the serious outcome of this cancer. Molecular & Cellular Proteomics

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confidence: 99%

Identification of Cellular Targets in Human Intrahepatic Cholangiocarcinoma Using Laser Microdissection and Accurate Mass and Time Tag Proteomics

Santos

Court

Thiers

et al. 2010

Molecular & Cellular Proteomics

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“…The utility of this information increases with the peak capacity of the separations and the reproducibility of peptide elution times [24,25]. Although the absolute LC elution time of a particular peptide can vary from run to run because of temperature and flow rate, among other factors, these changes can largely be corrected after normalization by using an appropriate algorithm to align multiple analyses [26,27].…”

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confidence: 99%

The Utility of Accurate Mass and LC Elution Time Information in the Analysis of Complex Proteomes

Norbeck

Monroe

Adkins

et al. 2005

J. Am. Soc. Mass Spectrom.

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The combination of mass and normalized elution time (NET) of a peptide identified by liquid chromatography-mass spectrometry (LC-MS) measurements can serve as a unique signature for that peptide. However, the specificity of an LC-MS measurement depends upon the complexity of the proteome (i.e., the number of possible peptides) and the accuracy of the LC-MS measurements. In this work, theoretical tryptic digests of all predicted proteins from the genomes of three organisms of varying complexity were evaluated for specificity. Accuracy of the LC-MS measurement of mass-NET pairs (on a 0 to 1.0 NET scale) was described by bivariate normal sampling distributions centered on the peptide signatures. Measurement accuracy (i.e., mass and NET standard deviations of Ϯ0.1, 1, 5, and 10 ppm, and Ϯ0.01 and 0.05, respectively) was varied to evaluate improvements in process quality. The spatially localized confidence score, a conditional probability of peptide uniqueness, formed the basis for the peptide identification. Application of this approach to organisms with comparatively small proteomes, such as Deinococcus radiodurans, shows that modest mass and elution time accuracies are generally adequate for confidently identifying most peptides. For more complex proteomes, more accurate measurements are required. However, the study suggests that the majority of proteins for even the human proteome should be identifiable with reasonable confidence by using LC-MS measurements with mass accuracies within Ϯ1 ppm and high efficiency separations having elution time measurements within Ϯ0.01

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“…Reduction of the complexity of protein mixtures can be reached at various levels. First, at the level of the tissue, methods of sample purification applied prior to analysis may well improve the accuracy of detection and increase the chance to identify low abundant proteins (18). By laser capture microdissection particular microscopic structures (like blood vessels) can be targeted and isolated and therefore reducing the chance of averaging out proteins of interest.…”

Section: Discussionmentioning

confidence: 99%

A Proteome Comparison Between Physiological Angiogenesis and Angiogenesis in Glioblastoma

Mustafa

Dekker²,

Stingl³

et al. 2012

Molecular & Cellular Proteomics

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The molecular pathways involved in neovascularization of regenerating tissues and tumor angiogenesis resemble each other. However, the regulatory mechanisms of neovascularization under neoplastic circumstances are unbalanced leading to abnormal protein expression patterns resulting in the formation of defective and often abortive tumor vessels. Because gliomas are among the most vascularized tumors, we compared the protein expression profiles of proliferating vessels in glioblastoma with those in tissues in which physiological angiogenesis takes place. By using a combination of laser microdissection and LTQ Orbitrap mass spectrometry comparisons of protein profiles were made. The approach yielded 29 and 12 differentially expressed proteins for glioblastoma and endometrium blood vessels, respectively. The aberrant expression of five proteins, i.e. periostin, tenascin-C, TGFbeta induced protein, integrin alpha-V, and laminin subunit beta-2 were validated by immunohistochemistry. In addition, pathway analysis of the differentially expressed proteins was performed and significant differences in the usage of angiogenic pathways were found. We conclude that there are essential differences in protein expression profiles between tumor and normal physiological angiogenesis. Molecular & Cellular Proteomics 11: 10.1074/mcp.M111.008466, 1-9, 2012.Neovascularization is a complex process taking place under physiological and pathological circumstances. There is large overlap in the cellular components, regulatory factors, and signaling mechanisms acting in the angiogenic process of regeneration, embryonic development, and tumor vascularization (1). In both physiological and pathological neovascularization signaling mechanisms, growth factors and their receptors, cell adhesion molecules and their specific extracellular matrix ligands take part (2). Angiocrine molecules like basic fibroblast growth factor and vascular endothelial growth factor and their receptors have been identified in the context of physiological and tumor angiogenesis (3). It is believed that the angiogenic switch is always triggered by hypoxia, starting with the up-regulation of vascular endothelial growth factor. The process of neovascularization consists not only of sprouting angiogenesis, but also of activation of endothelial precursor cells with the capacity to form de novo blood vessels (vasculogenesis). Vasculogenesis is also part of tumor vascularization but there is dispute about its relative importance (4, 5). Despite the similarities there are obvious differences between physiological vascularization and angiogenesis in tumors. Under physiological conditions, the regulatory mechanisms are well-coordinated and balanced and endothelial cell functions are tightly orchestrated by both pro-and anti-angiogenic factors. In tumor angiogenesis however, there is an excess of pro-angiogenic factors leading to uncoordinated proliferation and tubulogenesis of endothelial cells and migration of mural cells like pericytes (6). It is likely that the molecular diff...

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Nanoscale proteomics

Cited by 53 publications

References 24 publications

Identification of Cellular Targets in Human Intrahepatic Cholangiocarcinoma Using Laser Microdissection and Accurate Mass and Time Tag Proteomics

Identification of Cellular Targets in Human Intrahepatic Cholangiocarcinoma Using Laser Microdissection and Accurate Mass and Time Tag Proteomics

The Utility of Accurate Mass and LC Elution Time Information in the Analysis of Complex Proteomes

A Proteome Comparison Between Physiological Angiogenesis and Angiogenesis in Glioblastoma

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