Multiplexed electrokinetic sample fractionation, preconcentration and elution for proteomics.

Area of Science:

• Analytical Chemistry
• Microfluidics
• Proteomics

Background:

• Microfluidic devices are essential for complex sample analysis.
• Efficient sample pretreatment, including fractionation and preconcentration, is critical for sensitive detection.
• Existing methods often face challenges with reproducibility and analyte adsorption.

Purpose of the Study:

• To develop and characterize integrated microfluidic devices for "in space" sample fractionation, preconcentration, and elution.
• To create novel monolithic polymer beds with cationic properties for enhanced electroosmotic flow.
• To demonstrate the utility of these devices in a proteomics workflow.

Main Methods:

• Fabrication of 6 and 8-channel integrated microfluidic devices with monolithic porous polymer beds.
• Development of cationic surface coatings and polymer beds for anodal electroosmotic flow.
• Utilizing sheath flow assisted electrokinetic pumping for sample manipulation and elution.
• Characterization of elution reproducibility for neutral and charged analytes using fluorescence detection.

Main Results:

• The microfluidic devices successfully performed "in space" sample fractionation, preconcentration, and elution.
• A mixed cationic and hydrophobic monolithic polymer bed (META) provided effective sample capture and elution.
• Balanced electroosmotic flow prevented cross-contamination and reduced analyte adsorption.
• Reproducible elution performance was achieved with peak area reproducibility of ~8% and run-to-run reproducibility of 2.4-6.7% RSD.

Conclusions:

• The developed microfluidic devices offer a robust platform for integrated sample pretreatment in proteomics.
• The monolithic columns facilitate efficient elution and sheath flow generation.
• This technology is a key component for integrated fractionation and assay systems, enabling sensitive analysis by mass spectrometry.