References of "Massart, Anne-Cécile"
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See detailNovel post-digest isotope coded protein labeling method for phospho- and glycoproteome analysis
Fleron, Maximilien ULg; Greffe, Yannick ULg; Musmeci, Davide ULg et al

in Journal of Proteomics (2010), 73(10), 1986-2005

In the field of proteomics there is an apparent lack of reliable methodology for quantification of posttranslational modifications. Present study offers a novel post-digest ICPL quantification strategy ... [more ▼]

In the field of proteomics there is an apparent lack of reliable methodology for quantification of posttranslational modifications. Present study offers a novel post-digest ICPL quantification strategy directed towards characterization of phosphorylated and glycosylated proteins. The value of the method is demonstrated based on the comparison of two prostate related metastatic cell lines originating from two distinct metastasis sites (PC3 and LNCaP). The method consists of protein digestion, ICPL labeling, mixing of the samples, PTM enrichment and MS-analysis. Phosphorylated peptides were isolated using TiO(2), whereas the enrichment of glycosylated peptides was performed using hydrazide based chemistry. Isolated PTM peptides were analyzed along with non enriched sample using 2D-(SCX-RP)-Nano-HPLC-MS/MS instrumentation. Taken together the novel ICPL labeling method offered a significant improvement of the number of identified (∼600 individual proteins) and quantified proteins (>95%) in comparison to the classical ICPL method. The results were validated using alternative protein quantification strategies as well as label-free MS quantification method. On the biological level, the comparison of PC3 and LNCaP cells has shown specific modulation of proteins implicated in the fundamental process related to metastasis dissemination. Finally, a preliminary study involving clinically relevant autopsy cases reiterated the potential biological value of the discovered proteins. [less ▲]

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See detailNovel Relative ICPL Based Quantitative Phospho- and Glycoproteome Analysis Method
Fleron, Maximilien ULg; Greffe, Yannick ULg; Massart, Anne-Cécile ULg et al

Poster (2010, April 16)

Large scale proteomic analysis remains challenging partially because proteins are inhomogeneous and often influenced by a variety of structural modifications. In particular, these specific chemical ... [more ▼]

Large scale proteomic analysis remains challenging partially because proteins are inhomogeneous and often influenced by a variety of structural modifications. In particular, these specific chemical modifications called posttranslational modifications (PTM) are crucial determinants for the protein function and biological role. Up to now there have been a growing number of studies describing the enrichment and identification of PTM. However, a significant dearth of data offering a reliable methodology for PTM quantification does exist. The present work aims at developing a label based protein PTM quantification strategy and demonstrating its value on comparative analysis of cells originating from two distinct prostate metastasis sites. PC3 and LNCaP cells isolated from bone and lymph node prostate cancer metastasis sites respectively, were lysed and spiked with three non-human proteins serving as internal standards. Following this, the samples were reduced and alkylated, digested with trypsin and subjected to peptide ICPL (isotope coded protein label) labeling. The two peptide containing samples were joined together followed by the affinity isolation of phospho- (using TiO2 metal affinity chromatography) and glycopeptides (oxidized glycans were bound on hydrazide resin). The enriched fraction as well as the flow-through were analyzed on a 2D-(SCX and C18-RP)-nano-HPLC system. The peptide identification and quantification was conducted using electrospray ion-trap mass spectrometer (Bruker, HCT-ultra). Validation of the differentially modulated proteins was conducted in several biological and technical replicates using the label free MSe based quantification strategy. This PTM based, novel relative protein quantification using post-digest ICPL has detected over 598 individual proteins. Of these more than 95 % have been successfully quantified. PTM enrichment methodologies allowed an isolation rate of 91 % and 50 % for phosphorylated and glycosylated proteins respectively. The detailed comparison of PC3 and LNCaP cells has shown specific overexpression of selected proteins indicating differences between these two prostate metastatic cell lines. Several of these modulated proteins have been previously described to be related to prostate cancer (e.g. annexin A2 and vimentin) while others could be considered as potentially novel. These proteins might be implicated in the fundamental process related to metastasis dissemination. However, because of the known discrepancy between cell systems and clinical material, the present study can be regarded only as a step towards elucidation of these complex interactions. [less ▲]

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See detailMCF-7/BOS cells membrane proteome: comparison of two isolation methods using mass spectrometry
Bertrand, Virginie ULg; Massart, Anne-Cécile ULg; De Pauw, Marie-Claire ULg et al

Poster (2009)

Membrane proteins play an important role in biological processes but their isolation and quantification using classical techniques is often limited due to their poor solubility and relatively low ... [more ▼]

Membrane proteins play an important role in biological processes but their isolation and quantification using classical techniques is often limited due to their poor solubility and relatively low abundance. These membrane markers have to be accessible to antibodies and should be potential therapeutic targets. The tests were conducted on MCF-7 / BOS cell line, immortal and easier to cultivate. The goal of this work is to obtain a pure membrane fraction to facilitate the analysis of the sample. To isolate transmembrane proteins, we compared two methods. The first one used different extraction cycles characterized by different buffers to isolate membrane proteins. The second method labelled accessible extracellular domains at the surface of MCF-7 cells with biotin prior to differential centrifugation. [less ▲]

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See detailMCF-7/BOS cells membrane proteome: comparison of two isolation methods using mass spectrometry
Bertrand, Virginie ULg; Massart, Anne-Cécile ULg; De Pauw, Marie-Claire ULg et al

Poster (2009)

Membrane proteins (MP) play an important role in biological processes. Isolation and quantification of these MP using classical techniques is often limited due to their poor solubility and relatively low ... [more ▼]

Membrane proteins (MP) play an important role in biological processes. Isolation and quantification of these MP using classical techniques is often limited due to their poor solubility and relatively low abundance. These membrane proteins enclosed markers which could be potential therapeutic targets. These potential therapeutic targets have to be accessible to antibodies and need to be presented in the plasmic membrane. Assays were conducted on MCF-7 / BOS cell line, immortal and easier to cultivate. The goal of this work is to obtain an enriched membrane fraction to facilitate the analysis of the sample and to simplify the complex proteins mixture. To isolate transmembrane proteins, we compared two methods. The first one used different extraction cycles characterized by different buffers to isolate membrane proteins. The second method labelled accessible extracellular domains at the surface of MCF-7 cells with biotin prior to differential centrifugation. The obtained enriched membrane proteome was digested with trypsin and/or Lysyl Endopeptidase. Obtained peptides were separated by 2D-HPLC chromatography and on-line analysed ion trap mass spectrometer, the Esquire HCT. [less ▲]

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See detailDioxin-like compounds in porpoises and seals from the southern North Sea: relationship with biological and ecological factors
Das, Krishna ULg; De Pauw, Edwin ULg; Eppe, Gauthier ULg et al

in Organohalogen Compounds (2008), 70

The North Sea represents a major ecosystem for the harbour porpoise (Phocoena phocoena) and the harbour seal (Phoca vitulina). The grey seal (Halichoerus grypus) occurs more occasionally in the southern ... [more ▼]

The North Sea represents a major ecosystem for the harbour porpoise (Phocoena phocoena) and the harbour seal (Phoca vitulina). The grey seal (Halichoerus grypus) occurs more occasionally in the southern part of the North Sea. Their population over this last decade has experienced major fluctuations likely linked to prey availability and seal epizootics. Despite being banned more than 30 years ago, levels of polychlorinated biphenyls (PCBs) in marine mammals are still of concern due to historical contamination of the North Sea. [less ▲]

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See detailData quality of the serum analysis of PCDD, PCDF and PCB in the French dioxin and incinerators study
Pascal, M.; Eppe, Gauthier ULg; Zeghnoun, A. et al

in Organohalogen Compounds (2007), 69

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See detailHigh-throughput biomonitoring of dioxins and polychlorinated biphenyls at the sub-picogram level in human serum
Focant, Jean-François ULg; Eppe, Gauthier ULg; Massart, Anne-Cécile ULg et al

in Journal of Chromatography. A (2006), 1130(1), 97-107

We report on the use of a state-of-the-art method for the measurement of selected polychlorinated dibenzo-p-dioxins, polychlorinated dibenzofurans and polychlorinated biphenyls in human serum specimens ... [more ▼]

We report on the use of a state-of-the-art method for the measurement of selected polychlorinated dibenzo-p-dioxins, polychlorinated dibenzofurans and polychlorinated biphenyls in human serum specimens. The sample preparation procedure is based on manual small size solid-phase extraction (SPE) followed by automated clean-up and fractionation using multi-sorbent liquid chromatography columns. SPE cartridges and all clean-up columns are disposable. Samples are processed in batches of 20 units, including one blank control (BC) sample and one quality control (QC) sample. The analytical measurement is performed using gas chromatography coupled to isotope dilution high-resolution mass spectrometry. The sample throughput corresponds to one series of 20 samples per day, from sample reception to data quality cross-check and reporting, once the procedure has been started and series of samples keep being produced. Four analysts are required to ensure proper performances of the procedure. The entire procedure has been validated under International Organization for Standardization (ISO) 17025 criteria and further tested over more than 1500 unknown samples during various epidemiological studies. The method is further discussed in terms of reproducibility, efficiency and long-term stability regarding the 35 target analytes. Data related to quality control and limit of quantification (LOQ) calculations are also presented and discussed. (c) 2006 Elsevier B.V. All rights reserved. [less ▲]

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See detailStrategy for DR-CALUX dioxin screening in feed under EC regulation
Scippo, Marie-Louise ULg; Rybertt, M. S.; Eppe, Gauthier ULg et al

in Organohalogen Compounds (2006), 68

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See detailStrategy for DR-CALUX dioxin screening in feed under EC regulation
Scippo, Marie-Louise ULg; Rybertt, Soledad; Focant, Jean-François ULg et al

in Organohalogen Compounds (2006), 68

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See detailDioxin analysis in feed: cell-based assay versus mass spectrometry method
Scippo, Marie-Louise ULg; Rybertt, Soledad; Eppe, Gauthier ULg et al

in Accreditation and Quality Assurance (2006), 11(1-2), 38-43

In the determination of contaminants (dioxins, polychlorinated biphenyls, polyaromatic hydrocarbons), cell-based assays are useful methods for screening purposes: they are mainly characterized by high ... [more ▼]

In the determination of contaminants (dioxins, polychlorinated biphenyls, polyaromatic hydrocarbons), cell-based assays are useful methods for screening purposes: they are mainly characterized by high sample throughput and lower costs than the Mass Spectrometry (MS)-based methods. Although cell-based assays can be sensitive enough for the determination of dioxins and related substances in agreement with the presently tolerable limits in food and feed (Regulation No. 2375/2001/EC and Directive 2003/57/EC respectively), their lack of specificity make their use rather questionable in control laboratories. In this paper, we present and compare results obtained from the analysis of a limited number of feed samples by both gas chromatography-high resolution mass spectrometry (GC-HRMS) and cell-based assay (DR-CALUX: dioxin responsive-chemically activated luciferase gene expression) methods. The DR-CALUX screening led to less than 10% false non-compliant and no false compliant results. In addition, there is a good correlation between GC-HRMS and DR-CALUX data. However, these preliminary results have to be confirmed on a larger number of samples to demonstrate that total toxic equivalent (TEQ), including dioxins, furans and dioxin-like polychlorobiphenyls (PCBs) can be monitored in feed and food with a cell-based assay. [less ▲]

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See detailEnvironmental and human impact of an old-timer incinerator in terms of dioxin and PCB level: A case study
Pirard, Catherine; Eppe, Gauthier ULg; Massart, Anne-Cécile ULg et al

in Environmental Science & Technology (2005), 39(13), 4721-4728

The impact of a recently closed old municipal solid waste incinerator (MSWI) on polychlorodibenzo-p-dioxin (POD), polychlorodibenzofuran (PCDF), and polychlorinated biphenyl (PCB) levels in the ... [more ▼]

The impact of a recently closed old municipal solid waste incinerator (MSWI) on polychlorodibenzo-p-dioxin (POD), polychlorodibenzofuran (PCDF), and polychlorinated biphenyl (PCB) levels in the surrounding environment and resident serum has been studied in a small rural area of France. Studied soils and eggs from chickens foraging on these soils were sampled in the vicinity of the MSWI under the prevailing wind stream, while comparison samples were collected outside the assumed impact zone. PCB levels observed in soils and eggs did not differ statistically from comparison sites. This confirmed the low impact of MSWI PCB emission on environmental media, compared to other well-known sources. PCDD/PCDF levels in soils and eggs were significantly higher than in comparison samples, pointing out the impact of MSWI emission on the surrounding environment. The high dioxin concentrations in eggs set aside for private consumption would increase the dioxin intake for the studied population. Blood specimens of 10 nonoccupationally exposed volunteers who had lived within a 2 km radius of the incinerator for at least 25 years have been analyzed. When adjusted for age, PCB and PCDD/F blood levels were higher than general European populations and comparable to a similarly exposed Belgian population. [less ▲]

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See detailEvaluation of the DR-CALUX screening of food and feed, according to regulation levels including DL-PCB
Scippo, Marie-Louise ULg; Rybertt, Soledad; Focant, Jean-François ULg et al

in Organohalogen Compounds (2005), 67

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See detailComprehensive two-dimensional gas chromatography with isotope dilution time-of-flight mass spectrometry for the measurement of dioxins and polychlorinated biphenyls in foodstuffs - Comparison with other methods
Focant, Jean-François ULg; Eppe, Gauthier ULg; Scippo, Marie-Louise ULg et al

in Journal of Chromatography. A (2005), 1086(1-2), 45-60

A comprehensive two-dimensional gas chromatography time-of-flight mass spectrometry (GC x GC-TOF-MS) experimental setup was tested for the measurement of seven 2,3,7,8-substituted polychlorinated dibenzo ... [more ▼]

A comprehensive two-dimensional gas chromatography time-of-flight mass spectrometry (GC x GC-TOF-MS) experimental setup was tested for the measurement of seven 2,3,7,8-substituted polychlorinated dibenzo-p-dioxins (PCDDs), ten 2,3,7,8-substituted polychlorinated dibenzofurans (PCDFs), four non-ortho-polychlorinated biphenyls (PCBs), eight mono-ortho-PCBs, and six indicator PCBs (Aroclor 1260) in foodstuff samples. A 40 m RTX-500 (0.18 mm I.D., 0.10 mu m df) was used as the first dimension (D-1) and a 1.5 nn BPX-50 (0.10 mm I.D., 0.10 mu m df) as the second dimension (2 D). The GC x GC chromatographic separation was completed in 45 min. Quantification was performed using C-13-label isotope dilution (11)). Isotope ratios of the selected quantification ions were checked against theoretical values prior to peak assignment and quantification. The dynamic working range spanned three orders of magnitude. The lowest detectable amount of 2,3,7,8-TCDD was 0.2 pg. Fish, pork, and milk samples were considered. On a congener basis, the GC x GC-ID-TOF-MS method was compared to the reference GC-ID high resolution mass spectrometry (HRMS) method and to the alternative GC-ID tandem-in-time quadrupole ion storage mass spectrometry (QIST-MS/MS). PCB levels ranged from low picogram (pg) to low nanogram (ng) per gram of sample and data compared very well between the different methods. For all matrices, PCDD/Fs were at a low pg level (0.05-3 pg) on a fresh weight basis. Although congener profiles were accurately described, RSDs of GC x GC-ID-TOF-MS and GC-QIST-MS/MS were much higher than for GC-ID-HRMS, especially for low level pork and milk. On a toxic equivalent (TEQ) basis, all methods, including the dioxin-responsive chemically activated luciferase gene expression (DR-CALUX) assay, produced similar responses. A cost comparison is also presented. (c) 2005 Elsevier B.V. All rights reserved. [less ▲]

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See detailEvaluation of GC-MS/MS for determination of PBDEs in fish and shellfish samples
Pirard, Catherine; Eppe, Gauthier ULg; Massart, Anne-Cécile ULg et al

in Organohalogen Compounds (2005), 67

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