Zero-mode waveguides (ZMGs) are a new and powerful technique for reducing the volume of illuminated fluid in a fluorescence optical detection system. The method has been shown to allow unprecedented small observation volumes, resulting in new abilities in detecting and characterizing single molecules. Reduction of illuminated volume provides significant gains in signal-to-noise, signal-to-background and temporal resolution. Nanofluidics, Inc. intends to make these structures commercially available to industrial and academic users to enable research in and applications of single-molecule detection. One application with significant commercial potential has already been identified-- a single-molecule DNA sequencing strategy that requires a signal-to-noise that right now can only be attained with the ZMG technology. Other applications are expected to emerge when the research community is provided access the ZMGs. The performance of the ZMG has been proven, so the remaining technical risk parameters to commercialization are process consistency and production cost. The goals of this phase I program are to address the issues related to consistency in the production of ZMGs. First, it is essential to develop a tool to measure the interior dimensions of the ZMG, which is not possible with conventional inspection techniques. We propose to infer the dimensions of the structure from optical characteristics of the ZMG by comparison of optical measurements with simulation. We will use finite-element methods to simulate the electromagnetic fields in and around the ZMGs to tabulate both the fluorescence excitation profile and the efficiency of dipole radiation escape for a variety of geometries. Then a one-dimensional analytic diffusion model will be used to predict the fluorescence correlation spectra expected for each. Next, ZMGs will be fabricated and tested with the same geometries allowing validation of the models and measurement of fabrication variability. Success will be defined as demonstration of consistent attainability of sub 1-attoliter effective observation volumes, with variability in diameters of less than 25%. Upon completion of the phase I goals, Nanofluidics, Inc. will submit an SBIR phase II proposal under which we will make devices using the present methods and place them in beta test sites, while concurrently exploring methods of reducing the cost of manufacture. This exploration will range from incremental improvements of the process proven in Phase I to novel fabrication techniques that don't require expensive direct-write electron-beam lithography. [unreadable] [unreadable]