Fluorescent protein recognition polymer thin films capable of selective signal transduction of target binding events prepared by molecular imprinting with a post-imprinting treatment

Abstract

The functional monomer bearing three functional groups for protein imprinting was designed, which has a structure consisting of a polymerizable methacryloyl group, a secondary amine group for fluorescent dye conjugation by a post-imprinting treatment, and a benzoic acid moiety capable of interacting with a target protein. Lysozyme-imprinted polymer thin films were prepared on the initiator-immobilized glass substrates by radical polymerization in the presence of lysozyme, the designed functional monomer, a co-monomer(s) and a crosslinker. After the removal of lysozyme, fluorescein isothiocyanate was introduced into the secondary amine group of the functional monomer residues in the imprinted thin film as a fluorescent reporter dye (post-imprinting treatment). Lysozyme was selectively bound to the thin film with a binding constant of ca. 10(6) M(-1). Since the reporter dye can be only introduced into the binding cavity, the fluorescent response can be detected only when the guest is bound to the cavity, namely only specific binding events can be transduced as fluorescence spectral change. Compared with the SPR measurement, selective binding to the imprinted cavity can be more precisely detected by the proposed method, enabling us to prepare a new class of protein recognizable materials with the ability of the specific signal transduction of protein binding events.