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PXD006760

PXD006760 is an original dataset announced via ProteomeXchange.

Dataset Summary
TitleProteomic Sample Preparation in the Low Microgram Range – A Critical Evaluation of Current Workflows
DescriptionEfficient 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)).
HostingRepositoryPRIDE
AnnounceDate2018-05-12
AnnouncementXMLSubmission_2018-05-12_00:31:26.xml
DigitalObjectIdentifier
ReviewLevelPeer-reviewed dataset
DatasetOriginOriginal dataset
RepositorySupportUnsupported dataset by repository
PrimarySubmitterMalte Sielaff
SpeciesList scientific name: Mus musculus (Mouse); NCBI TaxID: 10090; scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationListmonohydroxylated residue; iodoacetamide derivatized residue
InstrumentSynapt MS
Dataset History
RevisionDatetimeStatusChangeLog Entry
02017-06-21 00:02:38ID requested
12017-10-02 04:15:05announced
22018-05-12 00:31:27announcedUpdated project metadata.
Publication List
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
curator keyword: Technical
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 affiliationInstitute for Immunology, University Medical Center of Mainz, Langenbeckstr. 1, 55131 Mainz, Germany
contact emailtenzer@uni-mainz.de
lab head
Malte Sielaff
contact affiliationInstitute for Immunology University Medical Center of the Johannes Gutenberg University Mainz
contact emailmalte.sielaff@gmail.com
dataset submitter
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