Large-area nanoplasmonic structures for enhanced fluorescence biosensing

Abstract: An innovative silver nano-dimple arrays (Ag DAs)-integrated microfluidic device was developed to achieve highly sensitive fluorescence-based nucleic acid detection. The Ag DAs were utilized to produce surface plasmon resonance (SPR) for strong fluorescence enhancement. We systematically investigated the plasmon-enhanced fluorescence by controlling the nanostructure dimension of the Ag DA substrate. The large-area periodic Ag DAs with controllable periodicity and dimple size were fabricated on a Si wafer by using nanosphere lithography (NSL), reactive ion etching (RIE), and Ag thin film deposition. The plasmonic behavior of the fabricated structures was studied both experimentally and theoretically using FDTD simulations. Fluorescence enhancement was observed to be more pronounced when the periodicity of the Ag DA was tuned to have its plasmonic resonance wavelength overlap the excitation and emission of fluorophores. An optimal enhancement factor of 172.565 was observed from the emission of fluorescein thin layer on the Ag DA of 400 nm periodicity. Fluorescence enhancement aside, the fluorophore in close proximity to the metallic surface of the Ag DA can be quenched efficiently. We exploited the distance-dependent fluorescence quenching and enhancement property to amplify the fluorescence response of the molecular beacons (MB) probes on Ag DAs. DNA sensing was conducted using the MB-immobilized Ag DA substrate integrated in a 1.71-µl microfluidic channel. The Ag DA yielded the maximum on-off fluorescence intensity ratio of 19.5 in response to the hybridization of target DNA, much larger than the ratio of 2.7 measured on a flat Ag surface The strong fluorescence enhancement of the Ag DA sensor yielded a low detection limit of 1 fM, equivalent to 1030 DNA copies in a 1.71-µl analyte volume. The high-performance DNA sensing technology shows promise for PCR-free point-of-care diagnostics applications.

Major Advisor: Li-Jing Cheng
Committee: Chih-hung Chang
Committee: John Labram
Committee: Matthew Johnston
GCR: Mei-Ching Lien