Single-Cell Secretome Profiling Enabled by Selective Enrichment and NanoLC-MS/MS Analysis
Yingyun He1,2, Xinxin Liu1, Fengjiao Zhu3,2
1State Key Laboratory of Medical Proteomics, National Chromatographic R. & A. Center, CAS Key Laboratory of Separation Science for Analytical Chemistry, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, 116023, China.
Angewandte Chemie (International Ed. in English)
|November 28, 2024
Summary
Researchers developed a novel single-cell secretome profiling method. This technique enhances the detection of low-abundance proteins, offering deeper insights into cellular communication and secretion heterogeneity.
Area of Science:
- Proteomics
- Cell Biology
- Biochemistry
Background:
- Single-cell proteomics allows profiling thousands of proteins from single cells.
- Investigating the single-cell secretome is challenging due to low-abundance proteins and high-abundance background proteins in serum-containing media.
- Existing methods struggle with comprehensive secretome analysis at the single-cell level.
Purpose of the Study:
- To develop a novel method for comprehensive single-cell secretome profiling (SCSP).
- To overcome the limitations of detecting low-abundance secreted and extracellular vesicle proteins.
- To enable deeper insights into secretion heterogeneity and intercellular communication.
Main Methods:
- Integration of metabolic labeling of newly synthesized proteins.
- Utilized click chemistry-based enrichment for protein isolation.
- Employed in situ digestion within an alkyne-functionalized capillary micro-reactor.
- Followed by nano liquid chromatography-tandem mass spectrometry (nanoLC-MS/MS) analysis.
Main Results:
- Quantified an average of 389 protein groups from the secretome of single HeLa cells (n=17).
- Confidently identified a total of 752 protein groups in the single-cell secretome.
- Achieved a significant increase in identified proteins compared to previous antibody-based targeted analyses.
Conclusions:
- The developed SCSP method provides a powerful tool for single-cell secretome analysis.
- Enables comprehensive investigation of secretion heterogeneity.
- Facilitates a deeper understanding of intercellular communication at the single-cell level.
