Limiting the Impact of Protein Leakage in Single-cell Proteomics

Overview

Journal bioRxiv

Date 2024 Aug 2

PMID 39091738

Authors

Andrew Leduc

Yanxin Xu

Gergana Shipkovenska

Zhixun Dou

Nikolai Slavov

Affiliations

Soon will be listed here.

Abstract

Limiting artifacts during sample preparation can significantly increase data quality in single-cell proteomics experiments. Towards this goal, we characterize the impact of protein leakage by analyzing thousands of primary single cells that were prepared either fresh immediately after dissociation or cryopreserved and prepared at a later date. We directly identify permeabilized cells and use the data to define a signature for protein leakage. We use this signature to build a classifier for identifying damaged cells that performs accurately across cell types and species.

References

Rosenberger F, Thielert M, Strauss M, Schweizer L, Ammar C, Madler S . Spatial single-cell mass spectrometry defines zonation of the hepatocyte proteome. Nat Methods. 2023; 20(10):1530-1536. PMC: 10555842. DOI: 10.1038/s41592-023-02007-6. View

Kelly R . Single-cell Proteomics: Progress and Prospects. Mol Cell Proteomics. 2020; 19(11):1739-1748. PMC: 7664119. DOI: 10.1074/mcp.R120.002234. View

Huffman R, Leduc A, Wichmann C, Gioia M, Borriello F, Specht H . Prioritized mass spectrometry increases the depth, sensitivity and data completeness of single-cell proteomics. Nat Methods. 2023; 20(5):714-722. PMC: 10172113. DOI: 10.1038/s41592-023-01830-1. View

Welch J, Kozareva V, Ferreira A, Vanderburg C, Martin C, Macosko E . Single-Cell Multi-omic Integration Compares and Contrasts Features of Brain Cell Identity. Cell. 2019; 177(7):1873-1887.e17. PMC: 6716797. DOI: 10.1016/j.cell.2019.05.006. View

Osorio D, Cai J . Systematic determination of the mitochondrial proportion in human and mice tissues for single-cell RNA-sequencing data quality control. Bioinformatics. 2020; 37(7):963-967. PMC: 8599307. DOI: 10.1093/bioinformatics/btaa751. View

Sanchez-Avila X, Truong T, Xie X, Webber K, Johnston S, Lin H . Easy and Accessible Workflow for Label-Free Single-Cell Proteomics. J Am Soc Mass Spectrom. 2023; 34(10):2374-2380. PMC: 11002963. DOI: 10.1021/jasms.3c00240. View

Schoof E, Furtwangler B, Uresin N, Rapin N, Savickas S, Gentil C . Quantitative single-cell proteomics as a tool to characterize cellular hierarchies. Nat Commun. 2021; 12(1):3341. PMC: 8185083. DOI: 10.1038/s41467-021-23667-y. View

Specht H, Emmott E, Petelski A, Huffman R, Perlman D, Serra M . Single-cell proteomic and transcriptomic analysis of macrophage heterogeneity using SCoPE2. Genome Biol. 2021; 22(1):50. PMC: 7839219. DOI: 10.1186/s13059-021-02267-5. View

Ilicic T, Kim J, Kolodziejczyk A, Bagger F, McCarthy D, Marioni J . Classification of low quality cells from single-cell RNA-seq data. Genome Biol. 2016; 17:29. PMC: 4758103. DOI: 10.1186/s13059-016-0888-1. View

10.

Massoni-Badosa R, Iacono G, Moutinho C, Kulis M, Palau N, Marchese D . Sampling time-dependent artifacts in single-cell genomics studies. Genome Biol. 2020; 21(1):112. PMC: 7212672. DOI: 10.1186/s13059-020-02032-0. View

11.

Lin B, Shah V, Chernoff C, Sun J, Shipkovenska G, Vinarsky V . Airway hillocks are injury-resistant reservoirs of unique plastic stem cells. Nature. 2024; 629(8013):869-877. PMC: 11890216. DOI: 10.1038/s41586-024-07377-1. View

12.

Gatto L, Aebersold R, Cox J, Demichev V, Derks J, Emmott E . Initial recommendations for performing, benchmarking and reporting single-cell proteomics experiments. Nat Methods. 2023; 20(3):375-386. PMC: 10130941. DOI: 10.1038/s41592-023-01785-3. View

13.

Leduc A, Khoury L, Cantlon J, Khan S, Slavov N . Massively parallel sample preparation for multiplexed single-cell proteomics using nPOP. Nat Protoc. 2024; 19(12):3750-3776. PMC: 11614709. DOI: 10.1038/s41596-024-01033-8. View