Targeting the Antibody Fab Region Using Light-Induced Indole-3-Butyric Acid Functionalized Magnetic Microspheres

Abstract

A novel light-controlled adsorption system for direct targeting of antibody Fab fragments was developed by utilizing indole-3-butyric acid functionalized magnetic microspheres. Indole-3-butyric acid, serving as a specific small molecule ligand, was successfully conjugated to amine-functionalized magnetic microspheres via a 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide activation strategy. Under illumination at a particular wavelength, the indole-3-butyric acid ligand generated reactive radicals that interacted with the nucleotide-binding sites of antibody Fab fragments, enabling effective affinity adsorption. Static adsorption experiments demonstrated that the system's adsorption behavior obeys the Langmuir model (K_F = 0.122, R² = 0.996), indicating a homogeneous adsorption process. Kinetic studies further revealed that the adsorption process follows a second-order kinetic model (k₂ = 0.0257, R² = 0.989). When compared with conventional antibody adsorption systems, this new system exhibited specific targeting of Fab fragments, enhanced selectivity, and adjustable properties. In particular, at pH 7.0, effective elution was achieved by increasing the salt concentration, with the eluted product retaining antigen-binding activity. The purification recovery rate exceeded 98%, and the system maintained effective adsorption and elution of Fab fragments across various pH conditions. Besides, even after 10 reuse cycles, the system retained more than 96% of its efficiency, thus presenting excellent regenerability and reusability. In summary, the developed light-controlled antibody Fab region adsorption system offers a highly efficient, stable, and cost-effective approach. It is also expected to become one of the most effective methods for antibody Fab purification in the future.