Toward immunoassay chips: Facile immobilization of antibodies on cyclic olefin copolymer substrates through pre-activated polymer adlayers

Abstract

Our research efforts have focused on the surface modification and the potential as a biochip material of commercially available cyclic olefin copolymer (COC) substrates. For the surface modification of hydrophobic COC substrates we synthesized two types of amphiphilic polymers having three important functions: hydrophobic (dodecyl or benzyl) groups, serving to anchor the COC substrate; a PEG component, which acts as a repellent of non-specific biomolecules; and an NHS ester group for conjugation of biomolecules. Formation of the polymer adlayers on COC surface was confirmed using a contact angle analyzer. The anti-biofouling property of the polymer-coated COC surface was examined by measuring the extent of nonspecific adsorption of immunoglobulin G (IgG), resulting in a very low level of the protein adsorption compared to uncoated COC surfaces (control). In addition, antibodies, used as representative biomolecules for immunoassay, could be selectively immobilized on the polymer-coated COC surface. Sandwich immunoassay using anti-rabbit IgG immobilized COC surface showed linear response to rabbit IgG of model analyte over a range from 1 to 1000 ng/mL with detection limit of 1-10 ng/mL. We also fabricated a Lab-On-a Chip type of COC biochip, which could detect a cardiac marker protein, troponin I (TnI), with detection limit of 10 ng/mL.