In this dissertation, I present signal processing techniques used in investigating single molecule kinetics and interactions. The first chapter provides an overview of signals and systems pertaining to single molecule studies. It discusses the need for signal processing, and gives a summary of current single molecule techniques. The second chapter introduces two techniques developed in our research using fluorescence microscopy, namely, spectral deconvolution in multi-color fluorescent microscopy systems; and fluorescent pattern recognition in the time domain using the correlation functions. In the third chapter, spectral deconvolution is demonstrated in separating autofluorescence from the fluorescence of enzyme label, used in quantitative fluorescence studies. In the fourth chapter, I discuss the fabrication of sub-wavelength metallic apertures that are used in single-molecule studies. I also discuss how optical properties of these devices are modeled and characterized. In the fifth chapter, I demonstrate how matched filter is used to give a precise quantitative measure of GFP fluorescence upon GFP aptamer binding. Similar technique is used to recognize photon bursts as a fluorescent molecule passes through an observation volume, discussed in the sixth chapter. I use this method to analyze the mobility of molecules in a fluidic channel. Chapter seven provides a summary and outlook for future studies.