The zeolitic imidazole framework with a leaf-shaped morphology (ZIF-L) has a wide range of promising applications in gas storage, battery materials, catalytic reactions, and optoelectronic devices due to its planar leaf-like structure and large surface area. However, the low conductivity, weak catalytic activity, and poor stability in the water dielectric medium of ZIF-L limit its further practical application. To solve these problems, we added the conductive polymer heterocyclic poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) (PEDOT:PSS) to ZIF-L for the sensitive detection of dopamine (DA). The synthesized composite ZIF-L/PEDOT:PSS (ZIF-L/PEDOT) not only retained the surface morphology of ZIF-L but also exhibited excellent electrochemical properties. The higher electrical conductivity of ZIF-L/PEDOT than that of ZIF-L was due to the enhanced electron transfer at the interface between ZIF-L and PEDOT:PSS. As a result, we developed an electrochemical biosensor based on the ZIF-L/PEDOT composite, which has a limit of detection of 7 nM for DA and a wide linear range from 25 nM to 500 μM. Furthermore, the current drop was negligible after 28 days, proving that the biosensor has excellent stability. Based on the above-mentioned outstanding performance, the ZIF-L/PEDOT-based biosensor was successfully used to detect DA in human serum samples. These results demonstrated that ZIF-L/PEDOT is expected to play an essential role in disease detection.