Methylcellulose enhances resolution in gravitational field-flow fractionation: Going beyond viscosity

Gravitational Field-Flow Fractionation (GrFFF) is an elution-based method designed for the separation of particles ranging from a few micrometers up to approximately 100 μm in diameter. Separation occurs over time, with particles being fractionated based on size and other physico-chemical properties. GrFFF takes advantage of gravitational forces acting perpendicularly to a laminar flow in a thin channel. The fluid exhibits a parabolic velocity profile, with the maximum velocity at the center of the channel and zero velocity at the walls. The exit time of particles depends on their equilibrium position relative to the bottom wall. In hyperlayer mode, larger particles elute faster than smaller ones due to their higher velocities within the channel. This study investigated the effect of adding methylcellulose (MC) to the carrier fluid on the elution behavior - specifically, peak time (t_peak) and resolution (R) - of polystyrene-based (PS) microparticles with sizes of 7, 8, and 10 μm. The results demonstrated that MC not only increases the viscosity of the carrier fluid but also exerts a secondary, predominant effect that improves resolution (R), thereby enhancing the separation of particle populations. This was confirmed by comparing the use of water as the carrier fluid at two different temperatures: 14 °C (high viscosity) and 28 °C (low viscosity). While increasing viscosity by lowering temperature only led to modest reduction in elution time of the fractograms, the addition of MC had a size-dependent effect on the microparticles, significantly improving R without changing other experimental parameters. This suggests the presence of additional phenomena contributing to the improved separation. In conclusion, the addition of MC to the carrier fluid increases the resolving power of GrFFF, enabling the separation of PS microbeads with a size difference of up to 2 μm. This advancement pushes the boundaries of GrFFF and opens up potential new applications. These studies, conducted on PS microbeads, provide a preliminary basis for future work on cells, which have similar density and size. This could pave the way for improved cell separation in diagnostic applications.