Semi-preparative asymmetrical flow field-flow fractionation: A closer look at channel dimensions and separation performance

Abstract

The design and performance of a semi-preparative asymmetrical flow field-flow fractionation (SP-AF4) channel are investigated with the objective of better understanding and exploiting the relationship between channel dimensions, sample loading, and resolution. Most size-based separations of nanometer and submicrometer particles are currently limited to analytical scale quantities (<100μg). However, there is a strong need to fractionate and collect larger quantities so that fundamental properties of the more narrowly dispersed fractions can be studied using additional characterization methods and for subsequent applications. In this work, dimensions of the spacer that defines the form of SP-AF4 channels are varied and their performances are assessed with respect to sample focusing position and loading. Separations are performed in aqueous and organic carrier fluids. A critical evaluation of channel dimensions showed that increasing the channel breadth is a practical and effective route to maintaining separation resolution while increasing sample loads to milligram quantities. Good size resolution (∼1.0) is achieved for separations of 10mg of 50 and 100nm silica nanoparticles suspended in water and up to 0.6mg of ∼10 to 35nm inorganic hybrid nanoparticles suspended in tetrahydrofuran. This work represents important advances in the understanding of SP-AF4 separations and extends sample loading capacities in both aqueous and organic solvents.

Recovery, overloading, and protein interactions in asymmetrical flow field-flow fractionation

“…The results demonstrate that not only the injected mass and the operational conditions are crucial, but also the type of the protein and presumably the solvent. Previous studies have investigated wider channels to increase sample loading [ 59 ]; we suggest that changes in the carrier liquid that aim to reduce the increase in viscosity with increasing sample concentration could also significantly increase the loading capacity. However, protein interactions are very complex, and therefore the factors that decrease viscosity are case specific.…”

Halophilic viruses with varying biochemical and biophysical properties are amenable to purification with asymmetrical flow field-flow fractionation

“…Injection volumes up to 1 L have been reported for dilute samples (Lee et al 1998). Alternative methods towards preparative AF4-purifications include increased channel dimensions (thicker or wider spacers) (Bria et al 2017). Sample dilution during AF4 is a known drawback of the method.…”

Asymmetrical flow field-flow fractionation for improved characterization of human plasma lipoproteins

“…[21] This, along with the unavailability of commercial MxHF5, led to the recent investigation of semi-preparative AF4 (SP-AF4) channel dimensions and performance. [27, 28] In comparison to analytical scale AF4, SP-AF4 uses larger channel breadths or thicknesses to increase the channel volume allowing for larger sample quantities to be introduced into the channel without overloading. Optimized SP-AF4 channels were able to handle milligram (up to 20 mg of 50 and 100 nm silica particles) quantities and are expected to provide an important capability for studies aimed at detection of low abundance biomarkers.…”