التفاصيل البيبلوغرافية
| العنوان: |
Automated single-cell proteomics providing sufficient proteome depth to study complex biology beyond cell type classifications. |
| المؤلفون: |
Ctortecka C; Broad Institute of MIT and Harvard, 415 Main Street, 02142 Cambridge, MA, USA., Clark NM; Broad Institute of MIT and Harvard, 415 Main Street, 02142 Cambridge, MA, USA., Boyle B; Broad Institute of MIT and Harvard, 415 Main Street, 02142 Cambridge, MA, USA., Seth A; Cellenion SASU, 60F avenue Rockefeller, 69008 Lyon, France., Mani DR; Broad Institute of MIT and Harvard, 415 Main Street, 02142 Cambridge, MA, USA., Udeshi ND; Broad Institute of MIT and Harvard, 415 Main Street, 02142 Cambridge, MA, USA., Carr SA; Broad Institute of MIT and Harvard, 415 Main Street, 02142 Cambridge, MA, USA. |
| المصدر: |
BioRxiv : the preprint server for biology [bioRxiv] 2024 Jan 22. Date of Electronic Publication: 2024 Jan 22. |
| نوع المنشور: |
Preprint |
| اللغة: |
English |
| بيانات الدورية: |
Country of Publication: United States NLM ID: 101680187 Publication Model: Electronic Cited Medium: Internet ISSN: 2692-8205 (Electronic) Linking ISSN: 26928205 NLM ISO Abbreviation: bioRxiv Subsets: PubMed not MEDLINE |
| مستخلص: |
Mass spectrometry (MS)-based single-cell proteomics (SCP) has gained massive attention as a viable complement to other single cell approaches. The rapid technological and computational advances in the field have pushed the boundaries of sensitivity and throughput. However, reproducible quantification of thousands of proteins within a single cell at reasonable proteome depth to characterize biological phenomena remains a challenge. To address some of those limitations we present a combination of fully automated single cell sample preparation utilizing a dedicated chip within the picolitre dispensing robot, the cellenONE. The proteoCHIP EVO 96 can be directly interfaced with the Evosep One chromatographic system for in-line desalting and highly reproducible separation with a throughput of 80 samples per day. This, in combination with the Bruker timsTOF MS instruments, demonstrates double the identifications without manual sample handling. Moreover, relative to standard high-performance liquid chromatography, the Evosep One separation provides further 2-fold improvement in protein identifications. The implementation of the newest generation timsTOF Ultra with our proteoCHIP EVO 96-based sample preparation workflow reproducibly identifies up to 4,000 proteins per single HEK-293T without a carrier or match-between runs. Our current SCP depth spans over 4 orders of magnitude and identifies over 50 biologically relevant ubiquitin ligases. We complement our highly reproducible single-cell proteomics workflow to profile hundreds of lipopolysaccharide (LPS)-perturbed THP-1 cells and identified key regulatory proteins involved in interleukin and interferon signaling. This study demonstrates that the proteoCHIP EVO 96-based SCP sample preparation with the timsTOF Ultra provides sufficient proteome depth to study complex biology beyond cell-type classifications.
Competing Interests: Conflict of interest A.S. is an employee of Cellenion. S.A.C. is a member of the scientific advisory boards of Kymera, PTM BioLabs, Seer and PrognomIQ. |
| التعليقات: |
Update in: Nat Commun. 2024 Jul 8;15(1):5707. doi: 10.1038/s41467-024-49651-w. (PMID: 38977691) |
| معلومات مُعتمدة: |
P01 CA206978 United States CA NCI NIH HHS; U01 CA271402 United States CA NCI NIH HHS; U24 CA270823 United States CA NCI NIH HHS; U24 CA271075 United States CA NCI NIH HHS |
| تواريخ الأحداث: |
Date Created: 20240208 Latest Revision: 20240722 |
| رمز التحديث: |
20240722 |
| مُعرف محوري في PubMed: |
PMC10849471 |
| DOI: |
10.1101/2024.01.20.576369 |
| PMID: |
38328197 |
| قاعدة البيانات: |
MEDLINE |
