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Related Experiment Video

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Laser Microdissection-Based Protocol for the LC-MS/MS Analysis of the Proteomic Profile of Neuromelanin Granules
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Integrated Proteome Analysis Device for Fast Single-Cell Protein Profiling.

Xi Shao1, Xuantang Wang1, Sheng Guan1

  • 1Department of Chemistry and Institutes of Biomedical Sciences , Fudan University , Shanghai 200433 , People's Republic of China.

Analytical Chemistry
|October 31, 2018
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Summary

A new integrated device, iPAD-1, enables single-cell proteome analysis within one hour. This high-sensitivity proteomic tool reveals significant cellular heterogeneity and aids in cell cycle phase estimation.

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Area of Science:

  • Proteomics
  • Single-cell analysis
  • Biotechnology

Background:

  • Previous work established an integrated proteome analysis device (iPAD-100) for analyzing proteomes from 100 cells.
  • Existing methods for single-cell proteome analysis are often time-consuming and suffer from sample loss.

Purpose of the Study:

  • To develop a novel integrated device (iPAD-1) for rapid and ultrasensitive single-cell proteome profiling.
  • To demonstrate the device's capability in analyzing protein expression heterogeneity and estimating cell cycle phases.

Main Methods:

  • A single cell is directly introduced into a capillary for simultaneous lysis and protein digestion in a minimal volume (2 nL).
  • Digestion is accelerated using elevated temperature and ultrasonication, with a total cell treatment time of 30 minutes.
  • Peptides are transferred to a nano-LC column with a 3 μm ESI tip for ultrasensitive detection using an Orbitrap Fusion Tribrid MS instrument.

Main Results:

  • The iPAD-1 successfully profiled proteins from single HeLa cells within 1 hour.
  • A maximum of 328 proteins were identified per cell, with a detection limit of 1.7-170 zmol.
  • The iPAD-1 demonstrated approximately 120-fold higher sensitivity compared to the iPAD-100 system.
  • Significant cellular heterogeneity in protein expression was observed.
  • Cell cycle phases were estimated based on core histone protein levels.

Conclusions:

  • The iPAD-1 represents a significant advancement in single-cell proteomic analysis, offering unprecedented speed and sensitivity.
  • The device effectively captures cellular heterogeneity, providing insights into individual cell functions.
  • This technology has potential applications in various fields, including cancer research and developmental biology.