Programmed-death ligand 1 (PD-L1), as one of major immune checkpoints, is highly expressed on cancer cells and participates in the immune escape process of tumor cells. The level of PD-L1 in patients is closely related to the efficacy of anti-PD-L1 immunotherapy, and patients with a high level have better response to immunotherapy. Therefore, PD-L1 can be an indicator of patient classification and medication guidance. In this work, we have developed a novel strategy for detecting PD-L1-positive circulating tumor cells based on steric hindrance generated after cell capture, using the primer exchange reaction (PER) amplification method. The principle is to modify a single strand containing the PD-L1 aptamer and the PER primer on the electrode surface. When PD-L1-positive circulating tumor cells exist, the aptamer will capture them. The steric hindrance generated by the captured cells due to their large volume hinders the subsequent approach of PER materials, thus hindering the occurrence of PER signal amplification. The number of HRP bound to the electrode surface is reduced, and the current signal output is inversely proportional to the number of captured cells. This method realizes convenient and sensitive detection of PD-L1-positive tumor cells and provides a new means for clinical judgment of whether patients should adopt immunotherapy.