Fabrication of macroporous microspheres with core-shell structure for negative chromatography purification of virus.

The macroporous microspheres with core-shell structure, based on a copolymer of 4-Vinylbenzyl chloride, glycidyl methacrylate, and ethylene glycol dimethacrylate, were fabricated through atom transfer radical polymerization suspension polymerization. The microspheres showed 100-200 nm pores in shell and 500-900 nm pores in core. The shell was hydrophilic modified through grafting of poly(N-hydroxyethyl acrylamide) onto the shell surface for reducing adsorption of proteins. The core was coupled with a ligand of poly(ethylene imine) that could bind the proteins. Feedstock of avian influenza virus could be purified on these modified microspheres through negative chromatography. Avian influenza virus cannot enter the core and was recovered from the flow-through, while other proteins with negative charges were able to penetrate into the core and bind to the poly(ethylene imine) ligands. The dynamic binding capacity of proteins was higher on this medium (61 mg/mL) than the commercially available resin (12 mg/mL, Capto Core 700).

Fabrication of poly(glycidyl methacrylate-co-ethylene glycol dimethacrylate) macroporous microspheres through activators regenerated by electron transfer atom transfer radical polymerization for rapid separation of proteins.

Activators regenerated by electron transfer atom transfer radical polymerization (AGET ATRP) were firstly used in suspension polymerization to prepare macroporous microspheres based on a copolymer of glycidyl methacrylate and ethylene glycol dimethacrylate. Compared to conventional radical polymerization (CRP), the microspheres by AGET ATRP showed more homogeneous structure, larger pores, and higher protein binding capacity. The body of microspheres are formed by the large clusters resulted from the aggregated little particles. The size of the particles in microspheres by AGET ATRP was 10-300 nm which was smaller than that (400-800 nm) of the microspheres by CRP. AGET ATRP gave larger pore size (275 ±â€¯5 nm) and surface area (59.3 ±â€¯1 m2/g) than CRP (234 ±â€¯5 nm, 37.5 ±â€¯1 m2/g). The microspheres were modified with polyethylene imine for anion resins that were evaluated in term of its protein binding capacity. The results indicated that the static (69 ±â€¯0.5 mg/mL) and dynamic binding capacity (61 ±â€¯0.5 mg/mL) of proteins on modified microspheres by AGET ATRP were higher than that (34 ±â€¯0.5 mg/mL and 19 ±â€¯0.5 mg/mL) by CRP. Meanwhile, the proteins binding capacity on the microspheres by AGET ATRP decreased only less than 10% when the flow rate increased 10 times. These macroporous media show a large potential in rapid separation of proteins.