PXD011689

PXD011689 is an original dataset announced via ProteomeXchange.

Title	CBB staining showed beneficial effects on proteomic analysis employing PAGE separation followed with whole-gel slicing, in-gel digestion and quantitative LC-MS/MS
Description	Proteomic analysis by combining PAGE separation, gel slicing and slice-by-slice LC-MS/MS has been frequently reported in recent years. Since the MS analysis would provide identities and quantities of the proteins along the whole lane, visualization by dye staining could be skipped to save time and labor and also in some reports assumed to improve MS identification and sensitivity. In this work, we examined the effect of CBB R-250 staining on the performance of the method and the results showed actually better results were obtained with CBB staining than without. A primary examination was firstly performed with gel bands of purified proteins, in which eight protein bands were excised, each from both the CBB-stained and unstained gel parts, and then analyzed by in-gel digestion and quantitative LC-MS/MS. Almost all the proteins were detected with higher sequence coverages and quantities from the stained gel bands than from the unstained. Then a proteomic sample of rat heart soluble proteins was examined for the complete workflow. The sample was firstly separated by nondenaturing PAGE and the gel was divided to two halves, with one CBB-stained and the other unstained. Laboratory-made tools were used to simultaneously cut duplicate lanes from each gel half and then slice each lane into about 39 pieces of the same size (1.1 mm 椋?1.1 mm 椋?1 mm thick). All the gel square pieces were analyzed in standardized procedures of in-gel digestion, peptide extraction and label-free quantitative LC-MS/MS. The results showed an average of 1434 proteins were detected in CBB-stained lanes, 40% higher than the 1013 in unstained lanes. When proteins detected in both conditions were compared, most of them were detected in higher quantities and in more gel squares with CBB staining. The comparison was also performed for SDS-PAGE and similarly advantageous results were obtained from the CBB-stained lanes, in both the detected numbers of proteins and peptides and the detected quantities and square numbers of individual proteins. The data also showed the proteins with lower molecular masses (e.g. < 30 kDa) were more benefited by the staining, probably because the dye binding helped to retain the proteins in gel matrix. In short, though dye staining is no longer a requisite when PAGE separation is followed by whole-gel LC-MS/MS analysis, CBB staining is still recommended for the better detection in proteomic analysis. All the raw and search files of the LC-MS/MS analysis, for (8*4=) 32 gel squares of HMW and LMW markers and (39*4+41*4=) 320 squares of rat heart soluble proteins (totally 704 files), as well as the gel patterns (2 files) and the summaries of the protein-level search results (3 files), are deposited in this project.
HostingRepository	PRIDE
AnnounceDate	2019-02-16
AnnouncementXML	Submission_2019-02-22_00:00:46.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Ya Jin
SpeciesList	scientific name: Rattus norvegicus (Rat); NCBI TaxID: 10116;
ModificationList	iodoacetamide derivatized residue
Instrument	Synapt MS

Revision	Datetime	Status	ChangeLog Entry
0	2018-11-13 08:10:20	ID requested
1	2019-02-15 21:57:00	announced
⏵ 2	2019-02-22 00:00:47	announced	Updated publication reference for PubMed record(s): 30776612.

Jin Y, Wen M, Yuan Q, Zhang J, Tan W, Beneficial effects of Coomassie staining on proteomic analysis employing PAGE separation followed with whole-gel slicing, in-gel digestion and quantitative LC-MS/MS. J Chromatogr B Analyt Technol Biomed Life Sci, 1110-1111():25-35(2019) [pubmed]

submitter keyword: CBB R-250 staining, nondenaturing PAGE, SDS-PAGE, proteomic analysis, whole gel slicing, in-gel digestion, quantitative LC-MS/MS

Ya Jin
contact affiliation	"Institute of Biomedical and Pharmaceutical Sciences, Guangdong University of Technology, Guangzhou 510006, China"
contact email	yajin@gdut.edu.cn
lab head
Ya Jin
contact affiliation	"Institute of Biomedical and Pharmaceutical Sciences, Guangdong University of Technology"
contact email	ya.jin@ymail.com
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2019/02/PXD011689

PRIDE project URI

• PRIDE
  1. PXD011689
     1. Label: PRIDE project
     2. Name: CBB staining showed beneficial effects on proteomic analysis employing PAGE separation followed with whole-gel slicing, in-gel digestion and quantitative LC-MS/MS