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In this study, a novel mercapto-terminated hexanuclear iron(III) cluster [Fe6O2(OH)2(O2CC6H4SCH3)10(hep)2]·CH3CN·CH2Cl2 [hep=2-(2-hydroxyethyl) pyridine] (Fe6) modified Au electrode was fabricated, having highly sensitive dopamine (DA) detection capabilities. In such Fe6 molecules, 10 thiomethyl groups are located at the periphery of the cluster, which enable the Fe6 molecules to self-assemble onto the surface of Au electrodes through the formation of Au–S bonds. The as-prepared Fe6-modified Au electrode (Au/Fe6) exhibits excellent electrocatalytic activity for the oxidation of dopamine (DA) in PBS with a diffusion coefficient of 3.12×10−5 cm2/s. Using the square wave voltammetry (SWV) technique, the calibration curve for DA determination was obtained in the range of 0.2 to 30 μM, and the detection limit for DA was ~0.07 μM. Furthermore, the modified electrode can accurately separate the DA signal from the interfering effect of uric acid (UA), thus providing simultaneous detection of DA and UA in their binary mixtures. This electrode can be reliably used to assay DA in its real drug composition.