Covalent and oriented immobilization of antibodies through systematic modification of photoactivatable RNA hybrid aptamers

Abstract

We report a covalent and well-oriented strategy of immobilization of antibodies using photoactivatable Fc-binding RNA aptamers (FcBAs). We prepared several types of FcBAs that were further modified with photoaffinity probes (i.e., benzophenone or diazirine), and evaluated the binding capabilities and the photo-crosslinking efficiency of them via pull-down assays and fluorescence analyses, respectively. Initial moderate photo-crosslinking efficiency (Kd ∼ 110 nM) was substantially improved by multivalent association of FcBAs and structural modification of FcBAs. For a conceptual proof, covalent crosslinking of human IgG on FcBA-tethered solid chips was fabricated and investigated by scanometry, which eventually proved real applicability of the present scheme toward immunoassays.