In this research, functionalized carbon nanotubes were employed to modify a glassy carbon electrode (F-MWCNT/GCE). Several techniques including Fourier Transform Infrared Spectroscopy, scanning electron microscopy, and transmission electron microscopy were utilized for characterization of F-MWCNT. The electrochemical activity of the developed electrode was assessed by differential pulse voltammetry, cyclic voltammetry, and electrochemical impedance spectroscopy. The electrochemical results demonstrated that oxidation of tryptophan (TRP) caused a higher anodic peak current than unmodified GCE. The F-MWCNT/GCE offered a detection limit of 3.63 nM and dynamic ranges of 0.01-0.7 µM for analysis of TRP by the differential pulse voltammetry method. Moreover, tryptophan detection was achieved successfully using F-MWCNT/ GCE in milk samples. It can be stated that this method paves the way for a selective, sensitive and straightforward method to determine tryptophan in the real food sample.