Novel ratiometric xanthene-based probes for protease detection

Novel FRET and donor-fluorescent quencher systems based on fluorescein-rhodamine B fluorophores have been developed as protease detectors and their applicability to chymotrypsin as a model protease sensing has been studied. The designed probes demonstrated long-term photostability and very good sensitivity in a chymotrypsin assay, allowing quantitative enzyme determination with the advantage of ratiometric measurement. The detection limit as well as linearity range for enzyme detection is tunable by the type of a linker between the dyes. The solid-phase synthetic protocol enables fast and easy adaptation of the system for various protease assays as well as its properties regulation.