Single-step covalent immobilization of oligonucleotides onto solid surface

Abstract

In this communication, we report a simple, single-step method for covalent immobilization of oligonucleotide probes onto an activated polystyrene surface. Polystyrene surface is activated by a photolinker, 1-fluoro-2-nitro-4-azidobenzene (FNAB) in a photochemical reaction by UV light. Amine-modified oligonucleotide probe is covalently immobilized to the activated surface by displacing the labile fluoro group of the activated polystyrene surface by the primary amino group of the aminated oligonucleotide probe. Biotinylated complementary oligonucleotide target is hybridized with the immobilized probe. The hybridized target is detected by binding streptavidin-peroxidase followed by assaying the enzyme for colorimetric signal measurement. The covalently immobilized oligonucleotide probe shows high selectivity in subsequent hybridization processes with the complementary target and clearly discriminates with single-base mismatched oligonucleotide targets. The method can immobilize amine- or thiol-modified oligonucleotide onto FNAB–activated polymer surface. The method could be potentially useful for designing DNA based biosensors or biochips and also in microtiter plate based assay systems in a cost effective manner.