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Herein, a cost-effective, simple and sensitive electrochemical sensing platform was established based on aptamer - target recognition and target-triggering signal amplification strategy for protein detection. Due to the high affinity between the aptamer and target, the assistant DNA1 (a1) could release from a1-aptamer duplex and trigger the following DNA circuits. The strand displacement and branch migration reaction brought assistant DNA3 (a3) released. Eventually, a large number of duplex structures of a3−Hairpin DNA3 were formed on the surface of electrode. Consequently, the capture DNA on the surface of platinum nanoparticles could hybridize with the unfolded DNA fragment of Hairpin DNA3 on the sensor surface, resulting in the electrochemical signal readout of H 2 O 2 reduction. Using thrombin as a model target, under the optimal conditions, this method exhibited a linear detection range from 0.5 pM to 300 nM with a detection limit of 0.17 pM. The proposed detection strategy was enzyme-free and exhibited good selectivity and sensitivity for a variety of protein targets detection by using corresponding DNA-based affinity probes, which makes it possible to apply the sensor for sensitivity detection of analytes in bioassays. A novel electrochemical sensing platform based on target-triggering cascade enzyme-free signal amplification strategy was developed for sensitive protein detection. [Display omitted] • Electrochemical detection of thrombin using target-triggering cascade enzyme-free signal amplification strategy was proposed. • The method was PCR-free, enzyme-free, low cost, and without the need for any additional separation steps. • The sensor for thrombin showed good performance. [ABSTRACT FROM AUTHOR]