High-performance monolith affinity chromatography for fast quantitation of immunoglobulin G

Abstract: High-performance monolith affinity chromatography employing protein A resins has been introduced previously for the fast purification of IgG from different sources. Here we describe the design and evaluation of a fast and specific method for quantitation of IgG from purified samples as well as crude supernatant from Chinese hamster ovary (CHO) cells. We used a commercially available affinity monolith with protein A as affinity ligand (CIM protein A HLD disk). Interferences of CHO host cell proteins with the quantitation of IgG from CHO supernatant were eliminated by a careful choice of the equilibration buffer. With this method developed, it is possible to quantify IgG within 5min in a concentration range of 23–250μg/ml. The calibration range of the method could be extended from 4 to 1000μg/ml by adjusting the injection volume. The method was successfully validated by measuring the low limit of detection and quantification, inter- and intra-day precision and selectivity. [Copyright &y& Elsevier]