Per- and polyfluoroalkyl substances (PFASs) are a class of persistent micropollutants. Due to their chemical stability and bioaccumulation, concentrations of PFASs in environmental media, even at ultratrace levels, pose significant environmental and health risks. However, currently reported detection methods lack an effective signal amplification strategy, and the detection sensitivity is limited, which can not meet the requirements of ultratrace detection. Herein, a groundbreaking aptamer-recognition-driven nucleic acid strategy was developed to significantly amplify the detection signal of perfluorooctanoic acid (PFOA). Furthermore, step pulse (SP) was used instead of cyclic voltammetry (CV) as an electrochemical excitation method to modulate the low electrochemiluminescence (ECL) triggering potential of poly [9,9-bis (3'-(N, N-dimethylamino) propyl) -2,7-fluorene]-alt-2,7-(9,9-dioctylfluorene)] (PFN) nanoparticles (NPs) so that a strong signal of +0.80 V was emitted without any exogenous coreactants. PFN NPs coupled rolling circle amplification-assisted PAM-free CRISPR/Cas12a system to construct an ultrasensitive ECL aptasensor for PFOA detection and the limit of detection was as low as 1.97 × 10-15 M. This ECL system integrated the advantages of no exogenous coreactants, low trigger potential, and nucleic acid amplification strategy and provided an ultrasensitive method for monitoring trace PFOA in the real water sample.