Sensitive, rapid, and accurate detection of dopamine (DA) at low cost is needed for clinical diagnostic and therapeutic purposes as well as to prevent illegal use of DA in animal feed. We employed a simple approach to synthesize reduced graphene oxide sheets (rGOS) and gold nanoparticles (AuNPs) at room temperature on indium tin oxide-coated glass (ITO) slides as disposable working electrodes for sensing DA. Graphene oxide (GO) was directly reduced on ITO to remove oxygenated species via a rapid and green process without using chemical reducing reagents. AuNPs were electrochemically deposited in situ on rGOS-ITO with fairly uniform density and size. The sensitivity of the AuNPs-rGOS-ITO sensor for DA detection is 62.7 μA mM⁻¹ cm⁻² with good selectivity against common electrochemically interfering species such as ascorbic acid (AA) and uric acid (UA), and the detection limit measured by differential pulse voltammetry (DPV), at a signal–noise ratio of 3, was 6.0 × 10⁻⁸ M. The electrochemical catalysis of DA was proven to be a surface process with an electron transfer coefficient (α) of 0.478 and a rate constant (k_s) of 1.456 s⁻¹. It correlates well with the conventional UV-vis spectrophotometric approach (R = 0.9973) but with more than thrice the dynamic range (up to 4.5 mM). The sensor also exhibited good stability and capability to detect DA in beef samples, and thus is a promising candidate for simple and inexpensive sub-nanomolar detection of DA, especially in the presence of UV-absorbing compounds.