Insulin sensors based on glassy carbon electrodes modified with nafion-multiwalled carbon nanotubes decorated with nickel hydroxide nanoparticles (Ni(OH)2NPs/Nafion-MWCNTs/GC), were prepared by electrochemical deposition of Ni(OH)2NPs from a dinuclear paddle-wheel Ni monothiocarboxylate complex on the MWCNTs/GC surface. The size and distribution of the Ni(OH)2NPs/Nafion-MWCNTs were characterized by transmission electron microscopy (TEM). The results show that Ni(OH)2 nanoparticles were electrodeposited on the surface of carbon nanotubes. Moreover, the electrochemical behavior of the modified electrodes in aqueous alkaline solutions of insulin was studied by cyclic voltammetry and chronoamperometry. It was found that the as-prepared nanoparticles have excellent electrocatalytic activity towards insulin oxidation due to their special properties, reducing the overpotential and improving the electrochemical behavior, compared to the bare GC electrode. Amperometry was used to evaluate the analytical performance of modified electrode in the determination of insulin. Excellent analytical features, including high sensitivity (5.0 A mol cm-2 μM-1), low detection limit (85 nM) and wide dynamic range (up to 10.00 μM), were achieved under optimum conditions. Moreover, these insulin sensors show good repeatability and a high stability after successive potential cycling. Common substances such as ascorbic acid, uric acid and acetaminophen do not interfere. Finally, the developed sensors have been applied to the determination of insulin in pharmaceuticals and in human plasma. Efficient recoveries for pharmaceuticals and human plasma demonstrate that the proposed methodology can be satisfactorily applied to these types of samples.