Thread-based isoelectric focusing coupled with desorption electrospray ionization mass spectrometry

Area of Science:

• Analytical Chemistry
• Biochemistry
• Mass Spectrometry

Background:

• Protein separation and preconcentration are crucial steps in mass spectrometry-based proteomics.
• Existing methods can be time-consuming, costly, and require complex instrumentation.
• Ambient mass spectrometry techniques like DESI-MS offer direct sample analysis but can benefit from improved sample preparation.

Purpose of the Study:

• To develop and validate a low-cost, integrated system for protein separation, concentration, and direct ambient mass spectrometry detection.
• To investigate the combination of thread-based isoelectric focusing (TB-IEF) with desorption electrospray ionization mass spectrometry (DESI-MS).
• To demonstrate the effectiveness of the combined platform for enhancing analyte response and matrix removal.

Main Methods:

• A 3D-printed thread-based isoelectric focusing (TB-IEF) device was fabricated and optimized for protein separation.
• Three model proteins were subjected to separation and focusing on the TB-IEF platform.
• The TB-IEF device was directly coupled with a DESI-MS system for analysis of the focused proteins.
• The impact of TB-IEF preconcentration on DESI-MS signal intensity and selectivity was evaluated.

Main Results:

• Successful separation and focusing of three model proteins were achieved within 30 minutes using the TB-IEF device.
• Direct coupling of TB-IEF with DESI-MS enabled "on-thread" ambient mass spectrometry detection.
• A 10-fold increase in DESI-MS response for insulin was observed after TB-IEF preconcentration.
• The method demonstrated simultaneous isolation of target proteins from the sample matrix.

Conclusions:

• The integrated TB-IEF and DESI-MS platform provides a novel, low-cost solution for protein sample pretreatment and focusing.
• This approach significantly enhances analyte detectability and selectivity in ambient mass spectrometry.
• The developed system offers a promising tool for rapid, on-site protein analysis with improved performance.