Enhancing purification efficiency of affinity functionalized composite agarose micro beads using Fe3O4 nanoparticles.

Abstract

In this work, a series of magnetic and nonmagnetic agarose matrices were fabricated for protein purification. Certain amounts of Fe3O4 nanoparticles were encapsulated in agarose beads to form composite magnetic matrices with enhanced purification efficiency. Structure and morphology of prepared matrices were studied by optical and scanning electron microscopes, FT-IR, and BET-BJH analysis. The prepared matrices had regular spherical shape, followed by a uniform size distribution. By nanoparticles addition, the number of mesopores decreased while population of pores with radius ≤10nm increased; thus, higher specific area achieved. According to VSM results, magnetization degree was one of the characteristics affected by agarose content of the beads. A dye ligand, Cibacron Blue F3GA (CB), was covalently bound to beads to adsorb Bovine serum albumin. CB concentration was determined by elemental analysis. It was shown that magnetic beads hold higher CB concentrations than nonmagnetic ones due to higher specific area. As a result, magnetic 8%-agarose beads had the highest affinity adsorption capacity in static experiments. Moreover, breakthrough curves were monitored to calculate dynamic binding capacity. And, it was shown that magnetic 4%-agarose had the highest adsorbing amount (6.00mg/mL). It was implied that pore diffusion in magnetic 4%-agarose may be the reason for higher dynamic capacity. Plus, column efficiency was evaluated. It was revealed that all magnetic beads had lower HETP (0.11, 0.12 and 0.11cm for magnetic 4, 6, and 8%-agarose beads) than nonmagnetic ones (P-value<0.05).