Dataset Information

Formaldehyde fixation helps preserve the proteome state during single-cell proteomics sample processing and analysis

ABSTRACT: Mass spectrometry-based single-cell proteomics (SCP) is gaining momentum but remains limited to a few laboratories due to the high costs and specialized expertise required. The ability to send samples to specialized core facilities would benefit non-specialist labs and popularize SCP for biological applications. However, no methods have been tested in SCP to "freeze" the proteome state while maintaining cell integrity for transfer between labs or prolonged sorting using fluorescence-activated cell sorting (FACS). This study evaluates whether short-term formaldehyde (FA) fixation can maintain cell states for shipping and on-site sorting, processing, and mass spectrometry analysis. We demonstrate that short-term FA fixation does not significantly affect protein recovery, even without heating and strong detergents, and maintains analytical depth compared to classical workflows. Fixation also preserves drug-induced specific perturbations of protein abundance during cell sorting and sample preparation for SCP analysis. Our findings suggest that formaldehyde FA fixation can facilitate SCP by enabling sample shipping and prolonged sorting, potentially democratizing access to SCP technology and expanding its application in biological research, thereby accelerating discoveries in cell biology and personalized medicine.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture, Hela Cell

SUBMITTER: Ilaria Piga

LAB HEAD: Jesper Velgaard Olsen

PROVIDER: PXD056327 | Pride | 2026-02-09

REPOSITORIES: Pride

ACCESS DATA

Dataset's files

Source:

			Action	DRS
	20240909_AST5_Evo3_IP_80SPD_4Th6ms_nDIA_Prot_OneTip_1000cells_1PC-FA_DMSO_1.raw	Raw
	20240909_AST5_Evo3_IP_80SPD_4Th6ms_nDIA_Prot_OneTip_1000cells_1PC-FA_DMSO_2.raw	Raw
	20240909_AST5_Evo3_IP_80SPD_4Th6ms_nDIA_Prot_OneTip_1000cells_1PC-FA_DMSO_3.raw	Raw
	20240909_AST5_Evo3_IP_80SPD_4Th6ms_nDIA_Prot_OneTip_1000cells_1PC-FA_DMSO_4.raw	Raw
	20240909_AST5_Evo3_IP_80SPD_4Th6ms_nDIA_Prot_OneTip_1000cells_1PC-FA_MTX_1.raw	Raw

Items per page:

1 - 5 of 143

Publications

Formaldehyde Fixation Helps Preserve the Proteome State during Single-Cell Proteomics Sample Processing and Analysis.

Piga Ilaria I Koenig Claire C Lechner Maico M Sabatier Pierre P Olsen Jesper V JV

Journal of proteome research 20250203 4

Mass spectrometry-based single-cell proteomics (SCP) is gaining momentum but remains limited to a few laboratories due to the high costs and specialized expertise required. The ability to send samples to specialized core facilities would benefit nonspecialist laboratories and popularize SCP for biological applications. However, no methods have been tested in SCP to "freeze" the proteome state while maintaining cell integrity for transfer between laboratories or prolonged sorting using fluorescen ...[more]

PMID: 39901769

Similar Datasets

Project description:Single-cell proteomics (SCP) has advanced significantly, yet it remains largely unidimensional, focusing primarily on protein abundances. This limitation hinders our understanding of the dynamic processes occurring within individual cells, particularly protein turnover, which is crucial for cellular function and regulation. In this study, we employed a pulsed stable isotope labeling by amino acids in cell culture (SILAC) approach to simultaneously evaluate protein abundance and turnover in single cells (SC-pSILAC). Using state-of-the-art SCP workflow, we demonstrated that two SILAC labels are detectable from ~4000 proteins in single HeLa cells recapitulating known biology. We investigated drug effects using protein synthesis and degradation inhibitors on global and specific protein turnover in single cells and performed a large-scale time-series SC-pSILAC analysis of undirected differentiation of human induced pluripotent stem cells (iPSC) encompassing six sampling times over two months and analyzed >1000 cells. Abundance measurements highlighted cell-specific markers of stem cells and various organ-specific cell types. Protein turnover dynamics highlighted differentiation-specific co-regulation of core members of protein complexes with core histone turnover discriminating dividing and non-dividing cells with potential in stem cell and cancer research. Lastly, correlating the abundance of individual proteins from cells displaying a wide range of diameters show that histones and some proteins involved in the cell cycle do not scale with cell size confirming previous observations in yeast. Our study represents the most comprehensive SCP analysis to date, offering new insights into cellular diversity and pioneering functional post-translational measurements beyond protein abundance. This method not only distinguishes SCP from other single-cell omics approaches and enhances its scientific relevance in biological research in a multidimensional manner but also showcase the discovery potential of SCP in fundamental biology.