Journal Article Epublish
PubMed: 40324992 DOI: 10.1038/s41467-025-56736-7
Limiting the impact of protein leakage in single-cell proteomics.
Nat Commun 16(1):4169
Authors: Andrew Leduc, Yanxin Xu, Gergana Shipkovenska, Zhixun Dou, Nikolai Slavov
Affiliations
- Departments of Bioengineering, Biology, Chemistry and Chemical Biology, Single Cell Proteomics Center, Northeastern University, Boston, MA, USA.
- Center for Regenerative Medicine, Massachusetts General Hospital, Boston, MA, USA.
- Departments of Bioengineering, Biology, Chemistry and Chemical Biology, Single Cell Proteomics Center, Northeastern University, Boston, MA, USA.
- Parallel Squared Technology Institute, Watertown, MA, USA.
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 from mouse trachea. The cells were prepared either fresh immediately after dissociation or first cryopreserved and prepared at a later date. We directly identify permeabilized cells by imaging a cell permeable dye and use the data to define a signature for protein leakage. This signature is similar across diverse cell types and reflects increased leakage propensities for cytosolic and nuclear proteins compared to membrane and mitochondrial proteins. A classifier based on the signature allowed for the accurate identification of permeabilized cells across cell types and species. The classifier is integrated into QuantQC ( scp.slavovlab.net/QuantQC ) to support its application to diverse samples and workflows.
MeSH
Cells, Proteomics