Rapid Purification of Immunoglobulin G Using a Protein A-​immobilized Monolithic Spin Column with Hydrophilic...

A rapid purification method was developed for antibody production in Chinese hamster ovary (CHO) cells using a Protein A-immobilized monolithic silica spin column with hydrophilic polymers. Monolithic silica modified with copolymers of 2-hydroxyethylmethacrylate (HEMA) and glycidyl methacrylate (GMA) showed lower non-specific protein absorption than that modified with a silane reagent. The epoxy group of GMA was converted to an amino group, and Protein A was modified by the coupling reagent. The amount of immobilized Protein A was controlled by changing the ratio of GMA to HEMA and the mesopore size of monolith. A modified monolith disk was fixed to a spin column for rapid antibody purification. The linear curves (for the antibody concentrations over 10 - 300 μg/mL) had a correlation coefficient of >0.999. Our column had various analytical advantages over previously reported columns, including a shorter preparation time (<10 min) and smaller sample volumes for purification with Protein A-immobilized agarose.