Modeling of protein electrophoresis in silica colloidal crystals having brush layers of polyacrylamide

Abstract

Sieving of proteins in silica colloidal crystals of mm dimensions is characterized for particle diameters of nominally 350 and 500 nm, where the colloidal crystals are chemically modified with a brush layer of polyacrylamide. A model is developed that relates the reduced electrophoretic mobility to the experimentally measurable porosity. The model fits the data with no adjustable parameters for the case of silica colloidal crystals packed in capillaries, for which independent measurements of the pore radii were made from flow data. The model also fits the data for electrophoresis in a highly ordered colloidal crystal formed in a channel, where the unknown pore radius was used as a fitting parameter. Plate heights as small as 0.4 μm point to the potential for miniaturized separations. Band broadening increases as the pore radius approaches the protein radius, indicating that the main contribution to broadening is the spatial heterogeneity of the pore radius. The results quantitatively support the notion that sieving occurs for proteins in silica colloidal crystals, and facilitate design of new separations that would benefit from miniaturization.

Comparison of separation modes for microchip electrophoresis of proteins

“…With the colloidal nanoparticle array, migration occurred in the order of increasing molecular mass, consistent with sieving. The apparent linear dependence of ln μ on molecular mass (Figure 3B) is consistent with free‐volume (Ogston) sieving models [12,17,25,26,45–47]. The model assumes that the ratio of electrophoretic mobility, μ , to the mobility in free solution μ 0 is equal to the fraction f of the total volume in a porous medium that can be occupied by the analyte during migration, given by [46–48]: …”

“…Silica colloidal crystal arrays afford an alternative separation mechanism in microdevices [1,12]. The colloidal array was formed in the separation channel of a PDMS chip with a glass base [1,12,25,26]. PDMS/glass chips were cast with wide channels (∼100 μm) from a mold with a thick PDMS layer (∼5 mm), resulting in relatively deep reservoirs.…”

“…Different sizes of nanoparticles create different pore sizes for size‐based separation of biomolecules [12]. The standard proteins were partially resolved with 400‐nm beads, which offer a pore size of ∼60 nm [12,26]. Separation was greatly improved with a 170 nm silica‐nanoparticle array (pore size, ∼26 nm) despite the short separation distance of ∼5 mm (Figure 3B).…”

Effect of Hydrophobic Moieties on the Assembly of Silica Particles into Colloidal Crystals

“…Since then, applications of CCs have noticeably increased in many fields; however, to date, this quote still applies to the field of separation science. Only few studies have reported the application of CCs as the media for microfluidic and chromatographic separation either in planar forms − or as packed, short microcapillaries (length ≤ 5 cm). − …”