We present a total internal reflection fluorescence microscopy (TIRFM) technique for quantitative molecular-level investigations of macromolecular adsorption and interactions at liquid-liquid interfaces. The technique provides the ability to selectively excite species within tens of nanometers of the interface and is an excellent tool for nonintrusive in situ investigations. The apparatus uses an oil-water assembly that is approximately 1.0 mm thick, enabling us to achieve a diffusion time constant of less than or equal to 3 min, an equilibrium adsorption time on the order of minutes rather than hours, and the possibility of using hydrodynamic shear to create new interfaces. In this paper, we give a detailed description of the apparatus and present some preliminary results of a study on the equilibrium adsorption of bovine serum albumin (BSA) and lysozyme at the oil-water interface.