<p> Leveraging the specific recognition capability of aptamers and the robust signal output of the Ru(bpy)₃²⁺/TPrA ECL system, a series of ECL-based aptasensors ha been developed for the detection of crucial small biomolecules. However, limitations such as low quenching efficiency and consequently limited sensitivity persist in current methods. To address these challenges, we have developed a low-background, high-output ECL signaling strategy and applied it to constructing a highly sensitive ECL aptasensor. Using ATP as a model target, the Fc-labeled aptamer effectively quenches the ECL signal, achieving a low background in the absence of ATP. Upon ATP binding, a conformational change in the aptamer restores and significantly enhances the ECL signal from the Ru(bpy)₃²⁺ luminophores. In this way, we have effectively integrated the aptamer, which serves as a specific recognition unit for the target, with a low-background, high-output ECL signaling system. The proposed sensor exhibited a linear response for ATP detection in the concentration range 10 pM to 1 µM, with a detection limit of 3.5 pM, along with excellent selectivity. The practical applicability and accuracy of the sensor were further demonstrated in real sample analyses. This method shows great promise for a wide range of applications in fields such as disease diagnosis and medical research.</p> Graphical abstract <p></p>