A self-assembling RNA aptamer-based graphene oxide sensor for the turn-on detection of theophylline in serum

To date, few effective fluorescent biosensors based on RNA aptamers have been developed because the intrinsic instability of RNA in the presence of nucleases precludes the application of RNA aptamers for the analysis of biological fluids. In this study, we developed a simple, sensitive, selective turn-on fluorescent aptasensor for theophylline detection in serum, utilizing ligand-induced self-assembling RNA aptamers and two different interaction stages of the aptamer fragments with graphene oxide (GO). A single strand of the theophylline RNA aptamer (33-mer) was split at the end loop region into two shorter fragments, one of which was labeled with a fluorophore (FAM). In the absence of theophylline, the adsorption of the two individual fragments on GO brought the fluorophore in close proximity to the GO surface, resulting in highly efficient quenching of fluorescence. The system showed very low background fluorescence. Conversely, the fragments self-assembled into an RNA aptamer/theophylline complex and were dissociated from GO. The quenched fluorescence was significantly recovered, and theophylline could be detected at a wide range of concentrations from 1 to 100μM, with a detection limit of 0.155μM and good selectivity in serum. Moreover, because of the shorter RNA fragments and the effective protection ability of GO from nuclease cleavage, the RNA sequences remained stable during the experiments. This design may serve as an example for the application of RNA aptasensors in the clinical setting.