PXD064954-1

PXD064954 is an original dataset announced via ProteomeXchange.

Title	Comparison of In-Gel and SP3 Mass Spectrometry Sample Preparation Protocols
Description	Sensitivity, robustness, and reproducibility of sample preparation are main determinants of data quality in bottom-up mass spectrometry-based proteomics. To this end, in-gel protein clean-up and digestion has been used for decades and is characterized by its robustness and compatibility with harsh lysis conditions. Single-pot solid-phase-enhanced sample preparation (SP3) has gained popularity recently and has been widely adapted as a standard workflow often replacing in-gel digestion-based workflows. Noteworthy, until today no direct comparison between both workflows has been made. Here, we performed a systematic comparison of in-gel and SP3 based sample preparation workflows assessing general performance, reproducibility, sensitivity, and fractionation possibilities on human cellular lysates and blood plasma. Both methods performed similarly regarding protein identifications, however, showed specific biases. SP3 outperformed the in-gel workflow regarding sample loss and thus demonstrated higher sensitivity when handling limited sample amounts, especially below 5 µg of input material. In contrast, in-gel sample preparation was superior in the identification of low molecular weight proteins. In conclusion, while SP3 is indeed an attractive state-of-the-art proteomics sample preparation method, in-gel digestion delivers competitive results and still has advantages in some applications, such as measurement of small proteins, e.g. in plasma samples.
HostingRepository	PRIDE
AnnounceDate	2026-08-24
AnnouncementXML	Submission_2026-08-23_20:06:05.985.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Dominic Helm
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: NEWT:9606;
ModificationList	iodoacetamide derivatized residue
Instrument	Orbitrap Exploris 480

Revision	Datetime	Status	ChangeLog Entry
0	2025-06-12 12:57:45	ID requested
⏵ 1	2026-08-23 20:06:06	announced

10.1002/pmic.70123;

Schwarzm, ü, ller L, Frey D, Zadora P, Schneider M, Wiemann S, Helm D, Comparison of In-Gel and SP3 Based Sample Preparation Protocols for LC-MS/MS Based Proteomics. Proteomics, 26(7):6-16(2026) [pubmed]

submitter keyword: None

Dr. Dominic
contact affiliation	Proteomics Core Facility Deutsches Krebsforschungszentrum Im Neuenheimer Feld 580 69120 Heidelberg Germany
contact email	d.helm@dkfz.de
lab head
Dominic Helm
contact affiliation	Proteomics Core Facility, German Cancer Research Center (DKFZ)
contact email	proteomicscore@dkfz.de
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD064954
     1. Label: PRIDE project
     2. Name: Comparison of In-Gel and SP3 Mass Spectrometry Sample Preparation Protocols