From Microsize Chromatographic Manufacturing for Fast Desalting to Its Characterization

Microfluidic devices are becoming increasingly popular in protein analysis due to their ability to reduce sample and buffer volumes. However, there is a research gap concerning the coupling of this technology with ion mobility and mass spectrometry (IM-MS). This study aims to fill this void by introducing the manufacture and the characterization of a microsize exclusion chromatography (μSEC) module for fast desalting and its integration into microfluidics, along with its coupling to electrospray ionization and ion mobility mass spectrometry (ESI-IM-MS). To assess the feasibility of this approach, the desalting of α-synuclein (αS) was investigated using Bio Spin P6 gel as a stationary phase in the manufacture of a microfluidic device. αS detection by MS gives insight into the sample purity, while IM combined with MS provides information about protein structure. IM allowed both the recording of qualitative and quantitative information. The qualitative data provided a map of the conformers in equilibrium, while the calculation of the respective abundances (quantitative profile) of the conformers afforded the opportunity to describe the dynamics of the system. Our experiments, serving as proof-of-concept, demonstrate αS desalting, exchange buffer efficiency, and reduced solvent usage, without compromising the protein's structure.