Surface modification of alumina monolithic columns with 3-aminopropyltetraethoxysilane (APTES) for protein attachment

Abstract

In the present work, a simpler and expedited functionalization method was used envisaging the preparation of functionalized alumina monoliths for effective purification of immunoglobulins. A single stage sol-gel synthesis method was used for the silanization of alumina monoliths with (3-Aminopropyl)triethoxysilane (APTES) in an aqueous environment. The nature of the attachment of APTES to the alumina and its distribution through the monolithic column was evaluated through FTIR-ATR, SEM-EDS and XPS measurements. Monolith silanization process was optimized by adjusting the reaction conditions in terms of catalyst used (acid or base), temperature and using a factorial design approach to elicit the interdependent influence of humidity, number of APTES coating layers and precursor concentration on the silanization of alumina. The reaction was found to be optimum at basic pH and a temperature of 80 °C. Etching-free functionalized monoliths with highest amine density of 166 µmol/g of the column were obtained using a single coating with a 2 M APTES solution, at 100% humidity. The binding capacity of the functionalized alumina monolith was determined using BSA and Protein A and then with the immunoglobulin G (IgG) aiming to infer about the suitability of the functionalized alumina monolith for purification of immunoglobulins. Adsorption-elution experiments showed that the functionalized alumina monoliths allowed the attachment of 0.78 mg IgG per mg of bound Protein A and the recovery of 89% of the captured IgG.