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Improved LC-MS/MS spectral counting statistics by recovering low-scoring spectra matched to confidently identified peptide sequences.

ABSTRACT: Spectral counting has become a popular method for LC-MS/MS based proteome quantification; however, this methodology is often not reliable when proteins are identified by a small number of spectra. Here, we present a simple strategy to improve spectral counting based quantification for low-abundance proteins by recovering low-quality or low-scoring spectra for confidently identified peptides. In this approach, stringent data filtering criteria were initially applied to achieve confident peptide identifications with low false discovery rate (e.g., < 1% at peptide level) after LC-MS/MS analysis and database search by SEQUEST. Then, all low-scoring MS/MS spectra that matched to this set of confidently identified peptides were recovered, leading to more than 20% increase of total identified spectra. The validity of these recovered spectra was assessed by the parent ion mass measurement error distribution, retention time distribution, and by comparing the individual low score and high score spectra that correspond to the same peptides. The results support that the recovered low-scoring spectra have similar confidence levels in peptide identifications as the spectra passing the initial stringent filter. The application of this strategy of recovering low-scoring spectra significantly improved the spectral count quantification statistics for low-abundance proteins, as illustrated in the identification of mouse brain region specific proteins.

SUBMITTER: Zhou JY

PROVIDER: S-EPMC2974764 | biostudies-literature | 2010 Nov

REPOSITORIES: biostudies-literature

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Improved LC-MS/MS spectral counting statistics by recovering low-scoring spectra matched to confidently identified peptide sequences.

Zhou Jian-Ying JY Schepmoes Athena A AA Zhang Xu X Moore Ronald J RJ Monroe Matthew E ME Lee Jung Hwa JH Camp David G DG Smith Richard D RD Qian Wei-Jun WJ

Journal of proteome research 20101004 11

Spectral counting has become a popular method for LC-MS/MS based proteome quantification; however, this methodology is often not reliable when proteins are identified by a small number of spectra. Here, we present a simple strategy to improve spectral counting based quantification for low-abundance proteins by recovering low-quality or low-scoring spectra for confidently identified peptides. In this approach, stringent data filtering criteria were initially applied to achieve confident peptide i ...[more]

PMID: 20812748

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Project description:ObjectiveThe aim of the current study was, first, to assess the coronary artery calcium (CAC) scoring potential of spectral photon-counting CT (SPCCT) in comparison with computed tomography (CT) for routine clinical protocols. Second, improved CAC detection and quantification at reduced slice thickness were assessed.MethodsRaw data was acquired and reconstructed with several combinations of reduced slice thickness and increasing strengths of iterative reconstruction (IR) for both CT systems with routine clinical CAC protocols for CT. Two CAC-containing cylindrical inserts, consisting of CAC of different densities and sizes, were placed in an anthropomorphic phantom. A specific CAC was detectable when 3 or more connected voxels exceeded the CAC scoring threshold of 130 Hounsfield units (HU). For all reconstructions, total CAC detectability was compared between both CT systems. Significant differences in CAC quantification (Agatston and volume scores) were assessed with Mann-Whitney U tests. Furthermore, volume scores were compared with the known CAC physical.ResultsCAC scores for routine clinical protocols were comparable between SPCCT and CT. SPCCT showed 34% and 4% higher detectability of CAC for the small and large phantom, respectively. At reduced slice thickness, CAC detection increased by 142% and 169% for CT and SPCCT, respectively. In comparison with CT, volume scores from SPCCT were more comparable with the physical volume of the CAC.ConclusionCAC scores using routine clinical protocols are comparable between conventional CT and SPCCT. The increased spatial resolution of SPCCT allows for increased detectability and more accurate CAC volume estimation.Key points• Coronary artery calcium scores using routine clinical protocols are comparable between conventional CT and spectral photon-counting CT. • In comparison with conventional CT, increased coronary artery calcium detectability was shown for spectral photon-counting CT due to increased spatial resolution. • Volumes scores were more accurately determined with spectral photon-counting CT.

Dataset Information

Improved LC-MS/MS spectral counting statistics by recovering low-scoring spectra matched to confidently identified peptide sequences.

Publications

Improved LC-MS/MS spectral counting statistics by recovering low-scoring spectra matched to confidently identified peptide sequences.

Similar Datasets

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