CE MALDI‐TOF/TOF MS for multiplexed quantification of proteins in human ventricular cerebrospinal fluid

Zuberovic, Aida; Wetterhall, Magnus; Hanrieder, Jörg; Bergquist, Jonas

doi:10.1002/elps.200800714

Cited by 33 publications

(26 citation statements)

References 40 publications

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“…Previously, a few groups have studied the proteome found in human serum [6, 7] or cerebrospinal fluid (CSF) [8, 11, 12] after TBI. These studies have focused on a general description of the proteome or selective study of unique proteins as predictive biomarkers.…”

Section: Discussionmentioning

confidence: 99%

Metabolic Crisis After Traumatic Brain Injury is Associated with a Novel Microdialysis Proteome

et al. 2010

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Background To examine if the metabolic distress after traumatic brain injury (TBI) is associated with a unique proteome. Methods Patients with severe TBI prospectively underwent cerebral microdialysis for the initial 96 h after injury. Hourly sampling of metabolism was performed and patients were categorized as having normal or abnormal metabolism as evidenced by the lactate/pyruvate ratio (LPR) threshold of 40. The microdialysate was frozen for proteomic batch processing retrospectively. We employed two different routes of proteomic techniques utilizing mass spectrometry (MS) and categorized as diagnostic and biomarker identification approaches. The diagnostic approach was aimed at finding a signature of MS peaks which can differentiate these two groups. We did this by enriching for intact peptides followed by MALDI-MS analysis. For the biomarker identification approach, we applied classical bottom-up (trypsin digestion followed by LC-MS/MS) proteomic methodologies. Results Five patients were studied, 3 of whom had abnormal metabolism and 2 who had normal metabolism. By comparison, the abnormal group had higher LPR (1609 ± 3691 vs. 15.5 ± 6.8, P < 0.001), higher glutamate (157 ± 84 vs. 1.8 ± 1.4 μM, P < 0.001), and lower glucose (0.27 ± 0.35 vs. 1.8 ± 1.1 mmol/l, P < 0.001). The abnormal group demonstrated 13 unique proteins as compared with the normal group in the microdialysate. These proteins consisted of cytoarchitectural proteins, as well as blood breakdown proteins, and a few mitochondrial proteins. A unique as yet to be characterized peptide was found at m/z (mass/charge) 4733.5, which may represent a novel biomarker of metabolic distress. Conclusion Metabolic distress after TBI is associated with a differential proteome that indicates cellular destruction during the acute phase of illness. This suggests that metabolic distress has immediate cellular consequences after TBI.

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Section: Discussionmentioning

confidence: 99%

Metabolic Crisis After Traumatic Brain Injury is Associated with a Novel Microdialysis Proteome

et al. 2010

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“…The iTRAQ technique identifies amino groups by using specific, isotope-code tags labelled peptides and tandem mass spectrometry analysis, stemmed from Applied Biosystems Inc (ABI) [18]. This technique is able to compare the relative levels of the same proteins from eight different samples simultaneously.…”

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confidence: 99%

Screening differential expression of serum proteins in AFP-negative HBV-related hepatocellular carcinoma using iTRAQ –MALDI-MS/MS

He¹,

Wang²,

W³

et al. 2014

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Hepatocellular carcinoma(HCC) is serious condition associated with a high morbidity and mortality. Therefore is an urgent need to develop novel noninvasive techniques for early diagnosis, particularly for patients with AFP-negative [AFP(-)] HCC. In this study, iTRAQ-MALDI-MS/MS was used to identify differentially expressed proteins in AFP(-) HBV-related HCC compared with non-cancerous hepatitis B virus (HBV) and healthy controls subjects.Serum was obtained from 18 patients with AFP(-) HBV-related HCC, 18 matched patients with HBV without HCC and 18 healthy control subjects. High abundance proteins were removed from serum and the differentially expressed proteins from the three groups were screened out using iTRAQ-MALDI-MS/MS. The Gene Ontology (GO) function and the interaction networks of differentially expressed proteins were then analyzed. A total of 24 expressed differential proteins associated with AFP(-) HBV-related HCC were screened out, 15 proteins were up-regulated and 9 down-regulated. The most common molecular function of the 24 differentially expressed proteins was enzyme inhibition. Interaction network of the 24 differentially expressed proteins showed that 14 proteins (C5, KNG1, FN1, LRG1, HRG, SERPINC1, CRP, APOB, SAA1, APCS, C4BPA, CFI, CFB and GSN) were central to the functional network. The expression levels of the GSN protein were down-regulated in AFP(-) HBV-related HCC subjects compared with healthy controls and the HBV group (p<0.01), consistent with the iTRAQ results.The 14 proteins from the serum of AFP(-) HBV-related HCC appeared at the fulcrum of the functional network and were differentially expressed compare to HBV and healthy controls suggesting a possible association with HCC progression.

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“…Capillary electrophoresis (CE), interfaced off-line to MALDI-TOF/TOF MS, was the first time used to monitor protein content in CSF with iTRAQTM labeling in patients with traumatic brain injury. [27] The analysis of ventricular CSF yielded to 43 significantly (p≤0.05 MudPIT scoring). Previously CSF has been characterized using a liquid phase isoelectric focusing in the Rotofor cell in combination with electroelution in the Mini whole cell gel eluter.…”

Section: Resultsmentioning

confidence: 99%

Robust Two-Dimensional Separation of Intact Proteins for Bottom-Up Tandem Mass Spectrometry of the Human CSF Proteome

et al. 2012

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The cerebrospinal fluid (CSF) is produced in the brain by cells in the choroid plexus at a rate of 500mL/day. It is the only body fluid in direct contact with the brain. Thus, any changes in the CSF composition will reflect pathological processes and make CSF a potential source of biomarkers for different disease states. Proteomics offers a comprehensive view of the proteins found in CSF. In this study, we use a recently developed non-gel based method of sample preparation of CSF followed by liquid chromatography high accuracy mass spectrometry (LC-MS) for MS and MS/MS analyses, allowing unambiguous identification of peptides/proteins. Gel-eluted liquid fraction entrapment electrophoresis (Gelfree) is used to separate a CSF complex protein mixture in 12 user-selectable liquid-phase molecular weight fractions. Using this high throughput workflow we have been able to separate CSF intact proteins over a broad mass range 3.5 kDa-100 kDa with high resolution between 15 kDa and 100 kDa in 2 hours and 40 min. We have completely eliminated albumin and were able to interrogate the low abundance CSF proteins in a highly reproducible manner from different CSF samples in the same time. Using LC-MS as a downstream analysis, we identified 368 proteins using MidiTrap G-10 desalting columns and 166 proteins (including 57 unique proteins) using Zeba spin columns with 5% false discovery rate (FDR). Prostaglandin D2 synthase, Chromogranin A, Apolipoprotein E, Chromogranin B, Secretogranin III, Cystatin C, VGF nerve growth factor, Cadherin 2 are a few of the proteins that were characterized. The Gelfree-LC-MS is a robust method for the analysis of the human proteome that we will use to develop biomarkers for several neurodegenerative diseases and to quantitate these markers using multiple reaction monitoring.

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CE MALDI‐TOF/TOF MS for multiplexed quantification of proteins in human ventricular cerebrospinal fluid

Cited by 33 publications

References 40 publications

Metabolic Crisis After Traumatic Brain Injury is Associated with a Novel Microdialysis Proteome

Metabolic Crisis After Traumatic Brain Injury is Associated with a Novel Microdialysis Proteome

Screening differential expression of serum proteins in AFP-negative HBV-related hepatocellular carcinoma using iTRAQ –MALDI-MS/MS

Robust Two-Dimensional Separation of Intact Proteins for Bottom-Up Tandem Mass Spectrometry of the Human CSF Proteome

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