Synthesis of Novel Copolymer for Selective Biomolecule Immobilization on Gold Surface

Abstract

Various types of nanoparticles such as quantum dots, magnetic nanoparticles, and liposomes have been studied for numerous biomedical applications. Among these nanoparticles, gold nanoparticles have been extensively exploited for various applications including drug delivery, imaging agents, biosensors, and computed tomography (CT) contrast agents because they have many advantages such as low cytotoxicity, easy size control, high x-ray absorption, unique optical and electrical properties. Stability of gold nanoparticles under physiological conditions is key step for biomedical application. Another key is the immobilization or conjugation of bioactive molecules on gold nanoparticles for targeted drug delivery, molecular imaging, and in vitro diagnosis. To satisfy these key requirements, we developed a novel random copolymer composed of a “surface anchor part” (thiol group), “antibiofouling part” (PEG), and “bioreactivity part” (carboxyl group) for gold surface modification in particular. A random polymer to modify the gold surface for targeting ligand conjugation or SPR (surface plasmon resonance) chip was synthesized using radical polymerization. The chemical structure of poly(SVE-r-mPEGMA-r-MA) is shown in Figure 1(a). This polymer consists of three parts: a S-4vinylbenzyl ethanethionate (SVE) monomer as the surfaceanchor part, poly(ethylene glycol) methyl ether methacrylate (mPEGMA) as the protein-resistant part, and methacrylic acid (MA) monomer as a functional group to conjugate biomolecules. The initial feed ratio (SVE: mPEGMA:MA, 2:2:1) was chosen for synthesizing polymer that have high antibiofouling effect and strong surface binding capacity. After radical polymerization, unreacted monomers were removed by dialysis. The structure of the polymer was confirmed using NMR. Peaks for the methoxy of the mPEGMA and the methyl of the SVE monomer were observed at 3.4 ppm and 2.3 ppm, respectively (Figure S1). The molecular weight of the polymer was Mn = 16 000, as measured by gel permeation chromatography (GPC). Next, we examined the ability of the polymer to coat the gold surface. First, the acetylated thiol of the polymer was deprotected using hydroxylamine to yield a free thiol group. Because several thiol groups in the polymer provide multipoint attachments to the gold surface, the polymer spontaneously forms monolayers on a gold surface. Because of the multiple attachments, the stability of the polymeric monolayers can be significantly improved when exposed to a buffer with a non-physiological pH, or oxidative chemical and electrochemical environments. A polymeric self-assembled monolayer (SAM) of the polymer on the gold surface was prepared by immersing the substrates in an ethanol solution of 0.2 wt % polymer. The polymer monolayer thickness was measured using ellipsometry. The ellipsometric thickness increased after immersion times of up to 4 h. After 4 h, the thickness of the films remained unchanged, and the thickness of the monolayer was approximately