In this work, the design and characterization of two fluorescent chemosensor dyes for two distinct analytes, respectively Naproxen and adenosine triphosphate (ATP), is fully described. For the first case (Naproxen chemosensor), the chemosensor dye consisted of styrylpyridinium fluorophore and the synthesis was performed in two reaction steps with relatively good yield. The sensing mechanism was based on twisted intramolecular charge transfer (TICT), with Naproxen inducing a fluorescence enhancement on the chemosensor. The limit of detection (LOD) for Naproxen was 0.054 mM. The sensor dye was used for the synthesis of polyacrylamide nanoparticles, PAA-NPs, with a diameter of 34 nm. The sensitivity of the PAA-NPs to Naproxen was decreased when compared to the free dye in solution with a LOD of 0.50 mM. Selectivity and competitive assays were performed and the obtained results indicate that the synthesized PAA-NPs can be used for rapid diagnosis of Naproxen overdose in human plasma samples. For the second case (ATP chemosensor), the sensor dye was based on a naphthalimide fluorophore and the synthesis comprised four reaction steps. The chemosensor presented a dipicolylamine unit able to complex with metal ions which allowed ATP binding, translated in a fluorescence enhancement of 2.3 fold. Results suggested that adenine appeared to present a preference in interaction with the naphthalimide fluorophore, probably through stacking interactions. The chemosensor was covalently immobilized onto amine surface-functionalized silica nanoparticles through NHS-ester activation. The obtained nanoparticles were found to have a hydrodynamic diameter of 126 nm. Their sensing properties towards ATP showed a significant decrease in sensitivity when compared to the dye in solution although sensing of concentrations in the low micromolar range was also found possible.