A Zwitterionic Detergent and Catalyst-Based Single-Cell Proteomics Using a Loss-Free Microhole-Collection Disc

Area of Science:

• Proteomics
• Mass Spectrometry
• Cell Biology

Background:

• Single-cell proteomics faces challenges in throughput, sample recovery, and scalability.
• Nanoscale sample handling is crucial for advancing single-cell analysis.
• Conventional mass spectrometry workflows often require specialized equipment for single-cell analysis.

Purpose of the Study:

• To develop a sensitive mass spectrometry (MS) analysis method for single cells using a conventional workflow.
• To overcome bottlenecks in throughput, sample recovery, and scalability in single-cell proteomics.
• To enable ultrasensitive proteomics analysis of limited samples without specialized nanovolume handling equipment.

Main Methods:

• Developed a mass-compatible single solution for seamless cell lysis and tandem mass tag (TMT) labeling using a zwitterionic detergent and catalyst.
• Utilized a micro-HOLe Disc (microHOLD) for single-cell lysis and TMT labeling without heat incubation.
• Collected TMT-labeled peptides through microholes in liquid chromatography injection vials in a single solution to prevent peptide loss.

Main Results:

• Achieved seamless cell lysis and TMT labeling in a single solution using the microHOLD device.
• Successfully applied the microHOLD method for analyzing proteome reprogramming in prostate cancer cells.
• Demonstrated high throughput and ultrasensitive proteomics analysis of single cells without specialized equipment.

Conclusions:

• The microHOLD method offers a novel, sensitive, and high-throughput approach for single-cell proteomics.
• This technique is not limited by the need for cutting-edge nanovolume handling equipment.
• The method facilitates the analysis of proteome reprogramming in limited samples, such as single cells, advancing cancer research.