The fluorescence and optical properties of membrane potential probes are widely used to measure cellular transmembrane potentials. Hemicyanine dyes are also able to bind to membranes. The spectral properties of these molecules depend upon the charge shift from the donor moiety to the acceptor moiety. Changes in their spectral properties, i.e. absorption and emission maxima or intensities, are helpful in characterizing model membranes, microheterogeneous media, etc. In this article, we have demonstrated the binding interaction of a membrane potential probe, 1-ethyl-2-(4-(p-dimethylaminophenyl)-1,3-butadienyl)-pyridinium perchlorate (LDS 698), with various supramolecular confined environments. The larger dipole moment in the ground state compared to the excited state is a unique feature of hemicyanine dyes. Due to this unique feature, red shifts in the absorption maxima are observed in hydrophobic environments, compared with bulk solvent. On addition of surfactants and CT DNA to an aqueous solution containing LDS 698, significant increase in the emission intensity along with the quantum yield and lifetime indicate partition of the probe molecules into organized assemblies. In the case of the sodium dodecyl sulfate (SDS)–water system, due to interactions between the cationic LDS 698 and the anionic dodecyl sulfate moiety, the fluorescence intensity at ∼666 nm decreases and an additional peak at ∼590 nm appears at premicellar concentration (∼0.20 mM–4.50 mM). But at ∼5.50 mM SDS concentration, the absorbance in the higher wavelength region increases again, indicating encapsulation of the probe in micellar aggregates. This observation indicates that the premicellar aggregation behavior of SDS can also be judged by observing the changes in the UV-vis and fluorescence spectral patterns. The temperature dependent study also indicates that non-radiative deactivation of the dye molecules is highly restricted in the DNA micro-environment, compared with micelles. Besides, we have also investigated the specific interaction of surfactant micelles with DNA. Our observations reveal that, in the presence of CT DNA, LDS 698 interacts exclusively with SDS micelles, but that it preferentially releases from micelles and relocates to DNA surfaces in solutions containing TX-100 micelles.