Fluorescent ionic liquids (FILs) incorporating the fluorescein anion have been prepared by anion exchange of the parent quaternary ammonium chloride (Quat(+)Cl(-)) ionic liquid. By controlling the molar ratio of fluorescein to Quat(+)Cl(-), ionic liquids incorporating different prototropic forms of fluorescein were prepared. The 1:1 molar ratio ionic liquid (FIL1) is essentially composed of monoanionic fluorescein, while dianionic fluorecein is predominant in the FIL with a 1:2 molar ratio (FIL2). The fluorescence excitation spectrum of FIL2 is markedly different from its absorption spectrum. Absorption features the fluorescein dianion, while the excitation spectrum is exclusively due to the monoanion. In FIL1, the absorption and excitation spectra are both characteristic of the monoanion. In both FILs, emission of the dianion is observed upon excitation of the monoanion. This unusual behavior is interpreted in the context of a fast deprotonation of the monoanion in the excited state. The presence of residual water in the ionic liquid is important for the proton transfer process. By lowering the pH of FIL1, the transient proton transfer is inhibited, and the emission of the monoanion could be observed. The FILs have completely different spectroscopic properties from solvated fluorescein in Quat(+)Cl(-), where the prototropic equilibrium is shifted toward the neutral forms.