Mass-Encoded Suspension Array for Multiplex Detection of Matrix Metalloproteinase Activities

Area of Science:

• Biomedical Engineering
• Analytical Chemistry
• Proteomics

Background:

• Matrix metalloproteinases (MMPs) play crucial roles in physiological and pathological processes.
• Accurate and sensitive detection of MMP activity is essential for disease diagnosis and drug development.
• Existing methods for multiplex MMP detection often face limitations in throughput, sensitivity, or specificity.

Purpose of the Study:

• To develop a novel mass spectrometric biosensing strategy for the multiplex detection of MMPs.
• To enable both relative and absolute quantification of MMP activity.
• To establish a high-throughput and ultra-sensitive platform for enzyme activity assays.

Main Methods:

• Fabrication of a mass-encoded suspension array using magnetic beads functionalized with MMP-specific peptide-isobaric tags for relative and absolute quantification (iTRAQ) conjugates.
• Integration of iTRAQ multiplex coding with ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS).
• Enzymatic reactions followed by trypsin digestion to release coding regions for UPLC-MS/MS analysis.

Main Results:

• Demonstrated multiplex detection of MMP-2 and MMP-7 with good linear responses in specified ranges (0.2-100 ng mL⁻¹ and 0.5-400 ng mL⁻¹, respectively).
• Achieved low detection limits of 0.064 ng mL⁻¹ for MMP-2 and 0.17 ng mL⁻¹ for MMP-7.
• Validated the strategy's specificity, practicability, and expansibility through analysis of MMP activity in serum samples and response to inhibitors.

Conclusions:

• The developed mass spectrometric biosensing strategy provides a powerful tool for multiplex enzyme activity assays.
• This approach offers high throughput, ultra-high sensitivity, and multiplexing capabilities for MMP detection.
• The work paves a new avenue for advancing mass spectrometric biosensing and enzyme activity profiling.