Localized surface plasmon-enhanced fluorescence spectroscopy for highly-sensitive real-time detection of DNA hybridization

Versatile and highly-sensitive detection of DNA hybridization is described using metal nanostructures-enhanced fluorescence (MEF) emission intensity when fluorescently-labeled DNA oligomers are covalently immobilized on a nanometer-thin amorphous silicon–carbon layer capping the metal nanostructures. The MEF structures are formed by thermal deposition of silver, gold or silver/gold thin films on glass surfaces and post-annealing at 500 °C. The choice of the metal film allows for tuning the optical properties of the interface. The metallic nanostructures are subsequently coated with an amorphous thin silicon–carbon alloy (a-Si0.80C0.20: H) layer deposited by PECVD. Carboxydecyl groups are attached on these surfaces through hydrosilylation then reacted with amine-terminated single-stranded DNA oligomers, forming a covalent link. The immobilized DNA is hybridized with its complementary strand carrying a fluorescent label. Through optimization of the thickness of the a-Si0.80C0.20: H alloy overlayer and by working close to resonance conditions for plasmon and fluorophore excitation, the hybridization of very dilute oligomers (5 fM) is easily detected, and the hybridization kinetics can be monitored in situ and in real-time.