Surface immobiliastion of antibody on cyclic olefin copolymer for sandwich immunoassay

There is a growing interest in the use of plastic materials in the development of biochips for various applications. In this work, the surface functionalisation of the commercially-available cyclic olefin co-polymer materials Zeonor (ZR) and Zeonex (ZX) was examined. The methodology developed entailed oxidation in oxygen plasma, functionalisation of the oxidized surface with aminopropyl triethoxy silane and attachment of antibodies using covalent linker molecules, namely 1,4-Phenylene diisothiocyanate (PDITC), Toluene-2,4-Diisocyanate and Sulfo-Succinimidyl-4-(N-Maleimidomethyl) cyclohexane-1-carboxylate. PDITC was selected as the most suitable cross-linker for the attachment of antibodies, as assessed by fluorescent intensity measurements on immobilised FITC-labelled IgG antibodies. The modification method was characterised by contact angle measurements, ellipsometry, x-ray photoelectron spectroscopy (XPS) and fluorescence microscopy. The data are consistent with the deposition of a polymeric film of APTES chemisorbed to the oxidised plastic surface. The functionalised surfaces were employed in a sandwich immunoassay format using the reagents goat anti-human IgG (GantiHIgG) and fluorescently-labelled GantiHIgG (Cy5-GantiHIgG) as capture and detection antibodies, respectively, and with human IgG (HIgG) as the model analyte. The lowest concentration of HIgG detected was 0.2 ng ml-1, with a relative standard deviation of 15%. Non-specific binding effects were also assessed. The method and supporting data demonstrate that simple approaches to surface functionalisation can be adapted to plastic chips.