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PXD006760

DataSet Summary

  • HostingRepository: PRIDE
  • AnnounceDate: 2018-05-12
  • AnnouncementXML: Submission_2018-05-12_00:31:26.xml
  • DigitalObjectIdentifier:
  • ReviewLevel: Peer-reviewed dataset
  • DatasetOrigin: Original data
  • RepositorySupport: Unsupported dataset by repository
  • PrimarySubmitter: Malte Sielaff
  • Title: Proteomic Sample Preparation in the Low Microgram Range – A Critical Evaluation of Current Workflows
  • Description: Efficient and reproducible sample preparation is a prerequisite for any robust and sensitive quantitative bottom-up proteomics workflow. Over the past years, several protocols been specifically developed for the preparation of samples, which are limited in quantity. In the present study, we carried out an independent comparison between three recently described methods: Single-Pot Solid-Phase-enhanced Sample Preparation (SP3), Filter Aided Sample Preparation (FASP) and a commercial kit based on the in-Stage Tip (iST) method. We assessed their performance for the processing of proteomic samples in the low μg-range using varying amounts of HeLa cell lysate (1 µg - 20 μg of total protein). All three workflows showed similar performance for 20 µg of starting material. When handling sample sizes below 10 µg, the number of identified proteins and peptides as well as the quantitative reproducibility and precision drastically dropped in case of FASP. In contrast, SP3 and iST provided high proteome coverage even in the low µg-range. Even when digesting 1 µg of starting material both methods still enabled the identification of over 3,000 proteins and between 25,000 to 30,000 peptides. On average, the quantitative reproducibility between experimental replicates was slightly higher in case of SP3 (R2= 0.97 (SP3); R2= 0.93 (iST)).
  • SpeciesList: scientific name: Mus musculus (Mouse); NCBI TaxID: 10090; scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
  • ModificationList: monohydroxylated residue; iodoacetamide derivatized residue
  • Instrument: Synapt MS

Dataset History

VersionDatetimeStatusChangeLog Entry
02017-06-21 00:02:38ID requested
12017-10-02 04:15:05announced
22018-05-12 00:31:27announcedUpdated project metadata.

Publication List

  1. Sielaff M, Kuharev J, Bohn T, Hahlbrock J, Bopp T, Tenzer S, Distler U, Evaluation of FASP, SP3, and iST Protocols for Proteomic Sample Preparation in the Low Microgram Range. J Proteome Res, 16(11):4060-4072(2017) [pubmed]

Keyword List

  1. curator keyword: Technical
  2. submitter keyword: Bottom-up proteomics, sample preparation, filter-aided sample preparation (FASP), single-pot solid-phase-enhanced sample preparation (SP3), in-StageTip digestion (iST)

Contact List

    Stefan Tenzer
    • contact affiliation: Institute for Immunology, University Medical Center of Mainz, Langenbeckstr. 1, 55131 Mainz, Germany
    • contact email: tenzer@uni-mainz.de
    • lab head:
    Malte Sielaff
    • contact affiliation: Institute for Immunology University Medical Center of the Johannes Gutenberg University Mainz
    • contact email: malte.sielaff@gmail.com
    • dataset submitter:

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