Integrating Low-Voltage Microfluidics With On-Chip Electrospray Ionization for Miniaturized Protein Analysis

Low-input proteomics requires analytical strategies that can handle very small volumes while minimizing loss, dilution, and handling steps. Low-voltage electrokinetic microfluidics offers a promising route to this goal by enabling controlled transport, separation, concentration, and droplet manipulation within compact chip-based platforms. This review examines the integration of electrokinetic microfluidics with on-chip electrospray ionization (ESI) for miniaturized protein analysis, with emphasis on how electric-field-driven sample handling can be combined with sensitive mass spectrometric detection. We discuss the fundamental electrokinetic mechanisms relevant to proteomic workflows, including electroosmotic flow (EOF), dielectrophoresis (DEP), electrowetting-on-dielectric (EWOD), electrohydrodynamic (EHD) flow, isotachophoresis (ITP), and electrothermal flow (ETF); and compare their operating conditions, strengths, and limitations for low-input analysis. We also review recent advances in on-chip ESI interfaces, emitter design, buffer management, and voltage-domain separation, highlighting the technical factors that govern stable coupling between microfluidic manipulation and mass spectrometry (MS) detection. Finally, we outline future directions in materials, automation, and intelligent workflow control that may improve robustness and reproducibility in next-generation electrokinetic proteomic platforms.