Specific binding of immunoglobulin G to protein A–mesoporous silica composites for affinity column chromatography

Abstract

High-performance affinity column chromatography is considered to be an important analytical tool for further advancements and popularisation of antibody–drug remedies. In this context, there is a great need for the design and development of a new protein A column system with desired functionalities at low cost. Here, we developed mesoporous silica (MPS) with pore diameters in the range of 2.3–31.2 nm, which was synthesised by a sol–gel method using organic templates, as a carrier for staphylococcal protein A. The surface of the MPS was organo-functionalised using γ-glycidoxypropyltrimethoxysilane (Gly), phenyltriethoxysilane (Ph) and 3-mercaptopropyltrimethoxysilane (SH). The specific binding of Immunoglobulin G (IgG) to the prepared protein A–MPS composites has been analysed as a function of pore size and surface properties. MPS composites with a smaller pore size (under 12.7 nm) demonstrate higher IgG binding activity than non-porous carrier composites. Of all the composites, the MPS-2.7–Gly–protein A composite, having mesochannels of size 2.7 nm and covalently bound to protein A, exhibits the highest binding efficiency. The molecular ratio of immobilised protein A to bound IgG is found to be approximately 1 : 1. These results imply that the composite MPS-2.7–Gly–protein A is at least 15 times more efficient than commercial carrier composites such as acrylic polymer–protein A. In addition, this composite shows remarkable stability towards low pH and chaotropic agents. Furthermore, the possibility of reusing the MPS-2.7–Gly–protein A composite is established by cycling it 5 times, with no loss in IgG extraction capacity. We expect that development of such MPS will have a significant impact on the industrial-scale advancement of high-efficiency and low-cost protein A affinity column chromatography.