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Ultra-high sensitivity mass spectrometry quantifies single-cell proteome changes upon perturbation

ABSTRACT: Single-cell technologies are revolutionizing biology but are today mainly limited to imaging and deep sequencing. However, proteins are the main drivers of cellular function and in-depth characterization of individual cells by mass spectrometry (MS)-based proteomics would thus be highly valuable and complementary. Here, we develop a robust workflow combining miniaturized sample preparation, very low flow-rate chromatography and a novel trapped ion mobility mass spectrometer, resulting in a more than ten-fold improved sensitivity. We precisely and robustly quantify proteomes and their changes in single, FACS-isolated cells. Arresting cells at defined stages of the cell cycle by drug treatment retrieves expected key regulators. Furthermore, it highlights potential novel ones and allows cell phase prediction. Comparing the variability in more than 430 single-cell proteomes to transcriptome data revealed a stable core proteome despite perturbation, while the transcriptome appears stochastic. Our technology can readily be applied to ultra-high sensitivity analyses of tissue material, posttranslational modifications and small molecule studies from small cell counts to gain unprecedented insights into cellular heterogeneity in health and disease.

INSTRUMENT(S): timsTOF

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Mario Oroshi

LAB HEAD: Matthias Mann

PROVIDER: PXD024043 | Pride | 2022-02-23

REPOSITORIES: Pride

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			Action	DRS
	DIANN1.8_SingleCells_CellCycle.7z	Other
	DIANN1.8_SingleCells_CellCycle.zip	Other
	DIANN1.8_diaPASEF_1ng_repetitions.zip	Other
	MQ_Dilutions_SingleCells_RawMinusMBR.zip	Other
	MQ_Dilutions_SingleCells_RawPlusMBR.zip	Other

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