AbstractMolecular imprinting is an advanced technology in the fabrication of novel chemical sensors recently applied, enabling rapid responses and highly selective binding to specific molecules, thereby enabling efficient detection of material constituents. In this study, a new molecularly imprinted electrochemical sensor (MIES) utilizing ferrocene‐functionalized carbon‐nanotubes (Fc‐CNTs) was developed for the determination of tryptophan (Trp). The formation of molecularly imprinted polymer (MIP) involves polymerization in dimethyl sulfoxide (DMSO) using Trp, methacrylic acid (MAA) and ethylene glycol dimethacrylate (EGDMA) as the template molecules, functional monomers, and cross‐linking agents, respectively. Due to the modification of MIPs, the sensor achieves high performance. The sensor demonstrates a linear range from 1 to 45 μM and from 45 to 330 μM, with the limit of detection of 0.44 μM, exceeding that of the majority of sensors reported in the literature. Furthermore, the sensor can effectively detect Trp in real samples, including human serum and amino acid oral liquid. The recovery rate of Trp in human serum samples is 94.2 % to 105.30 %, and in amino acid oral liquid is 94.3 % to 101.68 %. This study provides a promising approach for the effective detection and analysis of tryptophan constituents.