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The mechanism and kinetics of the electrochemical nucleation and growth of gold nanoparticles, AuNPs, onto a glassy carbon electrode, GCE, from Au(III) dissolved in the reline deep eutectic solvent, DES, at 70 °C, were assessed, for the first time. From the potentiodynamic technique it was found that gold electrodeposition onto the GCE substrate (Au(III)DES + 3e− (GCE) = Au(s)/GCE) is a diffusion-controlled process that requires of an overpotential nucleation to occur. The potentiostatic current density transients, recorded at different applied overpotentials, during the electrodeposition of AuNPs were described by a mechanism where multiple nucleation of 3D gold centers with mass-transfer controlled growth, occurs simultaneously with Au(III) adsorption (at the early stages) and the DES residual water reduction on the growing surfaces of the Au nuclei. From this analysis the diffusion coefficient of Au(III) ions was estimated as (2.56 ± 0.12) × 10−9 cm s−1 and it was found that the number density of Au nuclei, Ns, depends exponentially on the applied overpotential while the nucleation frequency, A, was practically constant and the water reduction contribution increases linearly. From SEM images and EDX spectrum of the GCE surface, electrodeposited with gold (GCE/AuNPs), it was found that the Au deposit was formed by aggregates, (183 ± 37) nm average size, of AuNPs (of ca 50 nm diameter) and a density of (1.8 ± 0.3) × 109 aggregates cm−2. The GCE/AuNPs was used for the Dopamine, DA, electrochemical quantification in the presence of uric acid, UA, with the following analytical performance: sensitivity of (32.49 ± 0.37) μA mM−1 and (28.6 ± 0.2) μM detection limit. [ABSTRACT FROM AUTHOR]