Petelski, Aleksandra A, Emmott, Edward 
ORCID: 0000-0002-3239-8178, Leduc, Andrew, Huffman, R Gray, Specht, Harrison, Perlman, David H and Slavov, Nikolai ORCID: 0000-0003-2035-1820
(2021)
Multiplexed single-cell proteomics using SCoPE2.
Nature protocols, 16 (12).
pp. 5398-5425.
ISSN 1754-2189, 1750-2799
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MultiplexedSCoPE2bioRxiv.pdf - Submitted version Download (2MB) | Preview |
Abstract
Many biological systems are composed of diverse single cells. This diversity necessitates functional and molecular single-cell analysis. Single-cell protein analysis has long relied on affinity reagents, but emerging mass-spectrometry methods (either label-free or multiplexed) have enabled quantifying >1,000 proteins per cell while simultaneously increasing the specificity of protein quantification. Here we describe the Single Cell ProtEomics (SCoPE2) protocol, which uses an isobaric carrier to enhance peptide sequence identification. Single cells are isolated by FACS or CellenONE into multiwell plates and lysed by Minimal ProteOmic sample Preparation (mPOP), and their peptides labeled by isobaric mass tags (TMT or TMTpro) for multiplexed analysis. SCoPE2 affords a cost-effective single-cell protein quantification that can be fully automated using widely available equipment and scaled to thousands of single cells. SCoPE2 uses inexpensive reagents and is applicable to any sample that can be processed to a single-cell suspension. The SCoPE2 workflow allows analyzing ~200 single cells per 24 h using only standard commercial equipment. We emphasize experimental steps and benchmarks required for achieving quantitative protein analysis.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | Oocytes, Hela Cells, U937 Cells, Animals, Humans, Mice, Peptides, Proteome, Indicators and Reagents, Chromatography, Liquid, Proteomics, Benchmarking, Tandem Mass Spectrometry, Single-Cell Analysis, Primary Cell Culture, RAW 264.7 Cells |
| Divisions: | Faculty of Health and Life Sciences Faculty of Health and Life Sciences > Institute of Systems, Molecular and Integrative Biology |
| Depositing User: | Symplectic Admin |
| Date Deposited: | 14 Mar 2022 16:35 |
| Last Modified: | 06 Dec 2024 18:58 |
| DOI: | 10.1038/s41596-021-00616-z |
| Related URLs: | |
| URI: | https://livrepository.liverpool.ac.uk/id/eprint/3150796 |
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