Interfacing microchip isoelectric focusing with on-chip electrospray ionization mass spectrometry

Area of Science:

• Analytical Chemistry
• Separation Science
• Microfluidics

Background:

• Microchip capillary isoelectric focusing (cIEF) offers miniaturized separation capabilities.
• Online mass spectrometric (MS) detection provides sensitive and specific analyte identification.
• Integrating these techniques on a single chip presents challenges in fluidic control and interface design.

Purpose of the Study:

• To develop and demonstrate a fully integrated microchip cIEF-ESI-MS system.
• To investigate a two-dimensional separation strategy combining cIEF and transient-isotachophoresis (tITP) for enhanced multiplexed separations.
• To evaluate the performance, selectivity, and speed of the developed microfluidic platform.

Main Methods:

• Fabrication of SU-8 microchannels for cIEF without surface pretreatment.
• Development of an on-chip sheath flow electrospray ionization (ESI) emitter for direct MS interfacing.
• Implementation of a two-dimensional chip for sequential cIEF and tITP separations.
• Optimization of electrokinetic mobilization using inherent electroosmotic flow (EOF).

Main Results:

• Successful interfacing of microchip cIEF with online MS detection via integrated ESI.
• Demonstration of a two-dimensional cIEF-tITP separation chip with improved selectivity beyond individual methods.
• Achieved short focusing times (60-270s) and total analysis times under 10 minutes.
• Repeatability of migration times for cIEF-tITP was 3.1-6.8% RSD (n=3).

Conclusions:

• The integrated microchip cIEF-ESI-MS system provides a powerful platform for rapid and selective analysis.
• The two-dimensional cIEF-tITP approach enhances separation power for complex samples.
• The use of native SU-8 channels and electrokinetic mobilization simplifies the system and reduces analysis time.