Going with the Flow: Mechanistic Insights into Slow Mixing Mode Native Mass Spectrometry

Area of Science:

• Analytical Chemistry
• Biophysical Chemistry
• Mass Spectrometry

Background:

• Slow mixing mode native mass spectrometry (SLOMO-nMS) is a powerful technique for in vitro quantification of biomolecular complexes.
• The method relies on mass balance principles, assuming constant analyte concentration during mixing within the nanoESI emitter.
• Deviations from mass balance can occur due to non-uniform mass transport in the emitter.

Purpose of the Study:

• To quantitatively investigate factors influencing solution mixing and analyte transport in nanoESI emitters under electric fields.
• To determine the relative contributions of diffusion, advection, and electrophoresis to analyte transport.
• To validate the mass balance assumptions crucial for SLOMO-nMS accuracy.

Main Methods:

• Utilized a dual-emitter setup with various dyes (varying size and charge) to study transport phenomena.
• Applied an electric field to the nanoESI emitter to mimic experimental conditions.
• Quantified diffusion, advection, and electrophoretic motion contributions to mixing and transport.

Main Results:

• Diffusion is identified as the primary driver for solution mixing and bulk transport.
• Advection becomes dominant at higher electric field voltages, influencing analyte transport.
• Electrophoretic displacement of analytes is found to be negligible at typical nanoESI voltages.
• The effective flow rate due to diffusion was quantified and found comparable to solution flow rates at low voltages.

Conclusions:

• Findings support the mass balance assumptions underlying SLOMO-nMS, enhancing its reliability.
• Understanding analyte transport mechanisms is critical for optimizing long-duration native mass spectrometry experiments.
• This research provides foundational insights for improving quantitative native MS techniques.