382187 Gigaporous Chromatography of Virus-like Particles
Virus-like particles (VLPs) represent a class of modern vaccines that are designed specifically for prevention of virus infections. These particles are produced through cultivation of recombinant microorganisms or mammalian cells. The cultivation suspension contains very low concentration of the product and large quantities of impurities. Hence it is necessary to purify the VLPs by chromatographic methods. Unfortunately, common chromatographic packing beads have pore sizes in the range of 20-30nm which close to the size of VLPs and may cause blockage of the pores. It is a prerequisite to develop suitable microsphere beads with much greater size, i.e. >100 nm to allow the VLPs to diffuse in and out of the beads freely. This study introduces the preparations of gigaporous microsphere beads which are made of polystyrene, poly-glycidylmethacrylate (PGMA) and agarose respectively. A simple method was developed for creating the super large pores, which was based on reverse micelles swelling. We found that surfactant reverse micelles could absorb water into the oil droplets, forming a bi-continuous W/S/O (water/surfactant/oil) emulsion. The aqueous channels became large pores after solidification of the oil droplets by polymerization. At the end, integral microspheres with giga-pores were obtained.
The effects of pore size on adsorption equilibrium, adsorption kinetics, dynamic binding capacity, and recovery were investigated. According to the confocal laser scanning microscopy, adsorption of VLPs in commercial chromatographic beads was mostly confined to a thin shell on the outer surface, leaving the underlying pore space and the binding sites inaccessibly, while the large pores in gigaporous beads enabled the VLPs to access to the interior pore spaces by diffusion transport efficiently. Ion exchange chromatography with the gigaporous beads gained about 12 times increase in static adsorption capacity, 8 times increase in dynamic binding capacity, and 11 times increase in effective pore diffusivity. The gigaporous structure also significantly improve the stability of the VLPs during intensive adsorption-desorption process by lowing the multi-point interaction between the VLPs and binding sites in the pores.