Highly efficient protein separations in capillary electrophoresis using a supported bilayer/diblock copolymer coating

Area of Science:

• Analytical Chemistry
• Separation Science
• Materials Science

Background:

• Capillary electrophoresis (CE) relies on stable electroosmotic flow (EOF) for reproducible separations.
• Surface modifications are crucial for controlling EOF and improving protein separation in CE.
• Traditional coatings often lack long-term stability under high-pressure buffer rinses.

Purpose of the Study:

• To develop and characterize a novel semi-permanent surfactant/polymer wall coating for capillary electrophoresis.
• To evaluate the stability and electroosmotic properties of the dimethyldioctadecylammonium bromide (DODAB) and polyoxyethylene (POE) 40 stearate coating.
• To assess the performance of the coating in separating diverse model protein mixtures.

Main Methods:

• A capillary was rinsed with a solution of DODAB and POE 40 stearate to form the wall coating.
• Electroosmotic flow (EOF) was measured and compared to coatings with DODAB alone.
• Model protein mixtures were separated across a pH range of 3-10.
• Separation efficiency, migration time reproducibility, and protein recovery were quantified.

Main Results:

• The DODAB/POE 40 stearate coating exhibited stable EOF, even after a 60-minute high-pressure buffer rinse.
• EOF was suppressed by over a factor of ten compared to DODAB-only coatings.
• High separation efficiencies (up to >1 million plates/m) were achieved for both basic and acidic proteins.
• Excellent migration time reproducibility (0.5-4.0% run-to-run, 0.6-4.3% day-to-day) and protein recoveries (84-97%) were observed.

Conclusions:

• The DODAB/POE 40 stearate coating provides a robust and reproducible platform for capillary electrophoresis.
• This semi-permanent coating significantly enhances protein separation performance in CE.
• The developed coating offers a promising alternative for analytical applications requiring stable and efficient protein separations.