Time-resolved fluorescence was applied to characterize the behavior in solution of a gemini surfactant substituted with pyrene (Py-3-12). Upon association in water, excimer formation by Py-3-12 can be probed by acquiring pyrene monomer and excimer fluorescence decays which can be fitted globally according to the model free (MF) analysis to yield quantitative information about the internal dynamics of the Py-3-12 surfactant micelles as well as a complete description of the distribution of the different pyrene species in solution either incorporated inside the micelles or free in solution. A proof of procedure for the MF analysis was established by noting that the concentrations of free surfactant in solution, [Py-3-12](free), was found to equal the critical micelle concentration (CMC) for surfactant concentrations larger than the CMC. (I-E/I-M)(SPC), the ratio of pyrene monomer to excimer fluorescence intensities, was calculated from parameters retrieved from the MF analysis of the fluorescence decays and was found to be independent of sample geometry. This work demonstrates how time-resolved fluorescence can be used to study the properties of pyrene-labeled macromolecules under conditions where large absorptions and inner filter effects usually distort the steady-state fluorescence signals. It was found that the pyrene excimer is formed mostly by diffusion within the Py-3-12 micelles, which suggests that the pyrene microenvironment is fluid, an important feature for future studies on the interactions of Py-3-12 with DNA.