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Related Experiment Video

Updated: Jun 23, 2025

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A Sample Preparation Procedure for Isobaric Labeling-Based Single-Cell Proteomics.

Consuelo Marín-Vicente1,2, Enrique Calvo3,4, José Manuel Rodríguez3

  • 1Cardiovascular Proteomics Group, Spanish National Centre for Cardiovascular Research (CNIC), Madrid, Spain. cmarin@cnic.es.

Methods in Molecular Biology (Clifton, N.J.)
|June 21, 2024
PubMed
Summary
This summary is machine-generated.

This study presents a method for single-cell proteomics using mass spectrometry. The protocol enhances sensitivity for analyzing limited biological samples, including difficult-to-isolate cells.

Keywords:
Mass spectrometryMultiplex sample preparation strategyOrbitrap Eclipse mass spectrometerSingle-cell proteomicsTandem mass tagTribrid mass spectrometer

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Area of Science:

  • Proteomics
  • Mass Spectrometry
  • Cell Biology

Background:

  • Traditional mass spectrometry-based proteomics requires substantial input material, limiting its application.
  • Single-cell proteomics demands ultra-high sensitivity due to minute sample quantities.
  • Existing methods face challenges in analyzing rare or difficult-to-isolate cell types.

Purpose of the Study:

  • To establish a general workflow for preparing cells for single-cell mass spectrometry-based proteomics.
  • To adapt isobaric labeling strategies with a carrier proteome to overcome sensitivity limitations.
  • To provide a protocol applicable to manually isolated cells, particularly large cells like cardiomyocytes.

Main Methods:

  • Utilizing isobaric labeling-based multiplex strategies.
  • Incorporating a carrier proteome to enhance signal detection.
  • Developing a general cell preparation workflow for mass spectrometry analysis.
  • Adapting the protocol for manual cell isolation techniques.

Main Results:

  • A robust protocol for single-cell mass spectrometry-based proteomics is demonstrated.
  • The method effectively addresses sensitivity limitations inherent in single-cell analysis.
  • The workflow is adaptable for challenging cell types, including those not amenable to standard sorting.

Conclusions:

  • The presented protocol significantly advances the feasibility of single-cell proteomics.
  • This approach broadens the scope of mass spectrometry applications in cell biology research.
  • The method offers a valuable tool for analyzing precious or rare cell populations.