Systematic optimization of targeted and multiplexed MS-based screening workflows for protein biomarkers

Area of Science:

• Biochemistry
• Analytical Chemistry
• Proteomics

Background:

• Targeted mass spectrometry (MS) enables sensitive, selective multiplexed measurement of biomarkers.
• Complex biological samples necessitate extensive preparation, hindering biomarker discovery.
• Current methods often require specific affinity reagents, limiting broad applicability.

Purpose of the Study:

• To develop and optimize a generic workflow for multiplexed protein biomarker screening.
• To eliminate the need for specific affinity reagents in the screening process.
• To enhance analytical performance through systematic optimization of sample processing and LC-MS methods.

Main Methods:

• Development of a surrogate peptide-based workflow for protein biomarker screening.
• Systematic assessment and optimization of each sample processing step.
• Optimization of liquid chromatography-mass spectrometry (LC-MS) parameters for enhanced performance.

Main Results:

• A generic, affinity reagent-free workflow for multiplexed screening of seven human proteins was established.
• Each stage of sample processing and LC-MS was optimized for improved analytical outcomes.
• The workflow demonstrated robust measurement capabilities for potential biomarkers.

Conclusions:

• The developed method facilitates efficient and sensitive protein biomarker screening.
• The workflow is applicable to complex biological samples, reducing preparation bottlenecks.
• The method was successfully applied to screen multiple myeloma patient samples for potential disease biomarkers.