Dataset Information

ThinPrep Cervical Smear iTRAQ LC-MS/MS

ABSTRACT: The protein content from ThinPrep cervical smear specimens was purified with TCA precipitation. The protein pellets were dissolved and protein concentration was measured with the Bradford assay. 100 ug of protein for each sample were reduced with TCEP, cysteines were blocked with MMTS and finally digested overnight with trypsin. The resultant peptides were labeled with the iTRAQ 8plex reagents and after mixing were subjected to off-line high pH C18 fractionation. The peptide fractions were further analyzed with low pH C18 LC-Orbitrap Elite HCD MS/MS (MS resolution 120,000, top 10 precursors, isolation width 1.2 m/z, 100 min gradient method). Protein Identification was performed using Proteome Discoverer 1.3 software. SEQUEST node included prec. mass tolerance: 10 ppm, fragment mass tolerance 0.5 Da, iTRAQ8plex Nterm and K,Y static modification, Methylthio static modification, Phospho S variable modification, Oxidation M variable modification, Deamidation N,Q variable modification. All spectra were search against a Uniprot fasta file containing all reviewed Human entries and all reviewed and un-reviewed human papillomavirus entries. Peptide and PTM validation was performed with Percolator and PhosphoRS nodes respectively. Protein quantitation was performed with the Reporter Ions Quantifier node.

INSTRUMENT(S): LTQ Orbitrap Elite

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Theodoros Roumeliotis

LAB HEAD: Theodoros Roumeliotis

PROVIDER: PXD000109 | Pride | 2013-05-09

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	02OCT12_cervical_smear_3rd_exp_20_rep.raw	Raw
	02OCT12_cervical_smear_3rd_exp_21_rep.raw	Raw
	02OCT12_cervical_smear_3rd_exp_22_rep.raw	Raw
	02OCT12_cervical_smear_3rd_exp_23_rep.raw	Raw
	02OCT12_cervical_smear_3rd_exp_24_rep.raw	Raw

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Publications

The shotgun proteomic study of the human ThinPrep cervical smear using iTRAQ mass-tagging and 2D LC-FT-Orbitrap-MS: the detection of the human papillomavirus at the protein level.

Papachristou Evaggelia K EK Roumeliotis Theodoros I TI Chrysagi Argyro A Trigoni Chrysanthi C Charvalos Ekatherina E Townsend Paul A PA Pavlakis Kitty K Garbis Spiros D SD

Journal of proteome research 20130422 5

The ThinPrep cervical smear is widely used in clinical practice for the cytological and molecular screening against abnormal cells and Human Papillomavirus (HPV) infection. Current advancements made to LC-MS proteomics include the use of stable isotope labeling for the in-depth analysis of proteins in complex clinical specimens. Such approaches have yet to be realized for ThinPrep clinical specimens. In this study, an LC-MS method based on isobaric (iTRAQ) labeling and high-resolution FT-Orbitra ...[more]

PMID: 23510160

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