A Simple and Flexible Strategy for Single-Cell Proteomic Analysis Based on Protein Immobilization and Digestion Tube...

Area of Science:

• Proteomics
• Biotechnology
• Analytical Chemistry

Background:

• Single-cell proteomics (SCP) analysis is rapidly advancing but hindered by complex, time-consuming preprocessing.
• Current methods require expensive equipment, specialized expertise, and multiple transfer steps, limiting widespread adoption.

Purpose of the Study:

• To develop a simplified, cost-effective, and rapid workflow for single-cell proteomics.
• To overcome the limitations of existing SCP preprocessing techniques.

Main Methods:

• A novel microliter reactor, the single-cell protein immobilization and digestion tube reactor (SPIDR), was engineered.
• SPIDR integrates cell sampling, protein preprocessing, and LC-MS/MS injection, eliminating additional transfer steps.
• The functionalized insert tube enables end-to-end processing within 1 hour.

Main Results:

• The SPIDR workflow quantified an average of 4186, 3171, and 4018 protein groups from single A549, HeLa, and MCF-7 cells, respectively.
• The microliter reactor design reduces operational difficulty and facilitates automation compared to nanoliter/picoliter systems.
• Demonstrated proteomic heterogeneity in cervical cancer cells undergoing apoptosis after paclitaxel treatment.

Conclusions:

• The SPIDR strategy offers a simple, flexible, and low-cost solution for rapid single-cell proteomics.
• This approach significantly enhances accessibility and efficiency for deep SCP analysis.
• SPIDR holds potential for advancing biological research by enabling detailed proteomic investigations at the single-cell level.