A label-free aptasensor for highly sensitive detection of ATP and thrombin based on metal-enhanced PicoGreen fluorescence

A label-free fluorescence aptasensor for highly selective and sensitive detection of ATP and thrombin was developed by using PicoGreen (PG) as signal molecule and surface-bound metal-enhanced fluorescence (MEF) substrates (silver island films, SIFs) as signal enhancers. On binding with ATP or thrombin, aptamers undergo structure switching, leading to a reduction of fluorescence intensity of PG. Chang of fluorescence intensity can be magnified by SIFs. The limit of detection for ATP and thrombin is 1.3 nM and 0.073 nM, respectively. The fluorescence quenching efficiency is linear in the logarithmic scale with ATP concentration range from 10 nM to 100 μM (R2=0.995) and thrombin concentration range from 0.1 nM to 100 nM (R2=0.997). The coefficients of variation of the intra-assay reproducibility and inter-assay reproducibility for ATP (10 μM) assay are 3.8% and 5.2%, respectively. In addition, the aptasensor is stable and can be reliably used for ATP measurement in biological samples. Overall, the aptasensor can be a useful and cost effective tool for the specific detection of ATP, thrombin and potentially other biomolecules in biological samples.