Three dimensional flower-like NiO-thermally reduced Graphene Oxide (trGO) nanocomposite was prepared by a simple hydrothermal method. The morphology and structure of the nanocomposite was characterized using Scanning Electron Microscope (SEM), Transmission Electron Microscope (TEM), Energy Dispersive X-ray (EDX), X-ray Diffraction (XRD) and Fourier Transform Infrared (FTIR) spectroscopy. The electrocatalytic oxidation of Dopamine, Ascorbic Acid (AA) and Uric Acid (UA) was investigated using Cyclic Voltammetry (CV). The modified electrode NiO-trGO/GCE showed good electrocatalytic activity compared to bare GCE. The detection of dopamine was carried out using Differential Pulse Voltammetry (DPV) wherefrom the Limit of Detection (LOD) and linear range was estimated to be 50 nM and 10 μM to 500 μM respectively. The selective detection of dopamine in presence of UA and AA was carried out. The NiO-trGO/GCE showed excellent selectivity for the detection of dopamine in presence of UA. However AA was found to interfere in the determination of dopamine. Also, the validity of the present sensor was examined in human biological samples.