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Glassy carbon electrodes (GCEs) modified with sulfur‐doped graphene (SG)/carboxylated carbon nanotube (CNT−COOH)/MoS2/yeast composite were prepared for electrochemical detection for lead ions by the simple hydrothermal methods and ultrasonic methods. The combination of SG and CNT−COOH could form a double‐layer carbon structure, providing more active detection sites for detection for lead, which could also contribute to adherence of yeast and MoS2. The SG/CNT−COOH/MoS2/yeast exhibited a high response in detecting low concentrations of lead ions. And then the SG/CNT−COOH/MoS2/yeast was characterized by Fourier transform infrared spectroscopy (FT‐IR), X‐ray diffraction (XRD), Transmission electron microscope (TEM), Scanning electron microscopy (SEM), and X‐ray photoelectron spectroscopy (XPS). Compared with traditional detection technology, the linear range of the sensor was 10−6∼10−14 g/L. And the lower of detection (LOD) down to 2.61×10−15 g/L was achieved. The sensor showed prospective applications in detection of Pb2+ in real serum samples.