Development and applications of condensed phase cavity ring-down spectroscopy for studies of electrochemical and interfacial.

Abstract

This dissertation reports the development of ultra-sensitive platforms based on the laser cavity ring-down spectroscopic (CRDS) technique to enable optical and spectroelectrochemical investigations in the condensed phase of matter at challenging scenarios. Firstly, an electrically-active solid/liquid interface for the evanescent-wave cavity ring-down spectroscopy (EW-CRDS) was developed to specroelectrochemically investigate redox events. By coating the interface of total internal reflection of the EW-CRDS platform with a high quality optically transparent and electrically conductive indium tin oxide thin film (ITO), we demonstrated that sufficiently long ring-down times can be achieved to allow for spectroelectrochemical investigations of redox species at solid/liquid interfaces at low surface coverages. The effects of an applied electric potential on the adsorption behavior of a redox protein onto different interfaces were investigated. For each interface, the adsorption and desorption constants, the surface equilibrium constant, the Gibbs free energy of adsorption, and the surface coverage were optically measured by our electrically-active EW-CRDS tool. Cyclic voltammetry (CV) scans under synchronous optical readout were performed to study the effects of each molecular interface in the redox process of surface-adsorbed protein species. The electro-active EW-CRDS technology is experimentally tested and demonstrated to provide a high-performance platform for studies of electrode-driven redox events of surface-confined molecular species at low submonolayer coverages and at a single diffraction-limited spot. Next, the electrically-active capability of the EW-CRDS device has been extended to develop a bio-sensing strategy based on the combination of the electro-active EW-CRDS platform with a sandwich immunoassay approach for the detection antigens of the influenza A virus (H5N1). Initially, the EW-CRDS was deployed to characterize in-situ and in real-time the formation of the assembly of the immunoassay-based biosensor. Our strategy proceeds in a stepwise manner: in the first step, the surface of the electro-active EW-CRDS device is functionalized with a capture antibody (Ab) aimed at a specific virus antigen. Next, the capture Ab-coated surface is exposed to a target antigen, which after binding to the surface it promotes the immobilization of secondary Ab that has been labeled with a redox-active probe. The redox-active probe methylene blue acts as a transduction element for monitoring molecular binding events and can be electrochemically modulated on the EW-CRDS platform to provide a unique optical interrogation signal. Based on this novel detection strategy, the experimental results have demonstrated an outstanding level of sensitivity in the pico-molar range for the detection of the influenza virus antigen. Finally, we used an electrically