PXD035339

PXD035339 is an original dataset announced via ProteomeXchange.

Title	On-capillary cell lysis enables top-down proteomic analysis of single HeLa cells by CE-MS/MS
Description	Proteomic analysis of limited samples and single cells requires specialized methods that prioritize high sensitivity and minimize sample loss. Consequently, sample preparation is one of the most important steps in limited sample analysis workflows to prevent sample loss. In this work, we have eliminated sample handling and transfer steps by processing intact cells directly in the separation capillary, online with capillary electrophoresis coupled to tandem mass spectrometry (CE-MS/MS) for top-down proteomic (TDP) analysis of low numbers of mammalian cancer cells (<10) and single cells. We assessed spray voltage injection of intact cells from a droplet of cell suspension (~1,000 cells) and demonstrated 0-9 intact cells injected with a dependency on the duration of spray voltage application. Application of spray voltage for 2 min injected an average of 7±2 cells and resulted in 33-57 protein and 40-88 proteoform identifications (N=4). To analyze single cells, manual cell loading by hydrodynamic pressure was used. Replicates of single cells (N=4) lysed on the capillary and analyzed by CE-MS/MS demonstrated a range of 17-40 proteins and 23-50 proteoforms identified. Furthermore, an additional cell line, THP-1, was analyzed at the single-cell level and proteoform abundances were compared to show the capabilities of single-cell top-down proteomics (SC-TDP) for assessing cellular heterogeneity. This study demonstrates the initial application of TDP in single-cell proteome-level profiling. These results represent the highest reported identifications from TDP analysis of a single HeLa cell and prove the tremendous potential for CE-MS/MS on-capillary sample processing for high sensitivity analysis of single cells and limited samples.
HostingRepository	PRIDE
AnnounceDate	2023-11-14
AnnouncementXML	Submission_2023-11-14_07:53:33.218.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Kendall Johnson
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	acetylated residue
Instrument	Orbitrap Fusion Lumos

Revision	Datetime	Status	ChangeLog Entry
0	2022-07-14 07:18:35	ID requested
1	2022-10-26 03:45:04	announced
⏵ 2	2023-11-14 07:53:34	announced	2023-11-14: Updated project metadata.

Johnson KR, Gao Y, Gregu, š M, Ivanov AR, On-capillary Cell Lysis Enables Top-down Proteomic Analysis of Single Mammalian Cells by CE-MS/MS. Anal Chem, 94(41):14358-14367(2022) [pubmed]

submitter keyword: top-down proteomics, single-cell top-down proteomics, tandem mass spectrometry, single-cell proteomics, capillary electrophoresis,CE-MS/MS

Alexander R. Ivanov
contact affiliation	Barnett Institute of Chemical and Biological Analysis, Department of Chemistry and Chemical Biology, Northeastern University, 360 Huntington Ave., Boston, Massachusetts 02115, United States
contact email	a.ivanov@northeastern.edu
lab head
Kendall Johnson
contact affiliation	Barnett Insititute of Chemical and Biological Analysis, Department of Chemistry and Chemical Biology, Northeastern University
contact email	johnson.kend@northeastern.edu
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/2022/10/PXD035339

PRIDE project URI

• PRIDE
  1. PXD035339
     1. Label: PRIDE project
     2. Name: On-capillary cell lysis enables top-down proteomic analysis of single HeLa cells by CE-MS/MS