The conformational flexibility of water-soluble p-(4-sulfonatophenyl) calixarene leads to pH dependent autofluorescence. Solution-based and solid-state techniques are consistent with the spontaneous formation of intra-molecular excimers upon solvation, with molecular simulations revealing that this occurs through π-stacking. Time-based fluorescence intensity studies provide insight into the solution dynamics, and solvated shelf-life, to significantly enhance the analytical protocol. Conductivity techniques support the formation of micelles, with no evidence for a direct link between higher-order structures and autofluorescence. Disruption of this excimer by divalent metal cations provides scope for its utility as a fluoroionophore, which is also consistent with molecular simulations, without the requirement of incorporating additional fluorophoric and water solubilising sub-units.