PXD035339 is an
original dataset announced via ProteomeXchange.
Dataset Summary
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 | 2022-10-26 |
AnnouncementXML | Submission_2022-10-26_03:45:03.988.xml |
DigitalObjectIdentifier | |
ReviewLevel | Peer-reviewed dataset |
DatasetOrigin | Original dataset |
RepositorySupport | Unsupported dataset by repository |
PrimarySubmitter | KendallJohnson |
SpeciesList | scientific name: Homo sapiens (Human); NCBI TaxID: 9606; |
ModificationList | acetylated residue |
Instrument | Orbitrap Fusion Lumos |
Dataset History
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. |
Publication List
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] |
Keyword List
submitter keyword: top-down proteomics, single-cell top-down proteomics, tandem mass spectrometry, single-cell proteomics, capillary electrophoresis,CE-MS/MS |
Contact List
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 | |
KendallJohnson |
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 | |
Full Dataset Link List
Dataset FTP location
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PRIDE project URI |
Repository Record List
[ + ]
[ - ]
- PRIDE
- PXD035339
- Label: PRIDE project
- Name: On-capillary cell lysis enables top-down proteomic analysis of single HeLa cells by CE-MS/MS