Silver nanoparticles (nanoAg) incorporated poly(3,4-ethylene dioxythiophene-sodium dodecyl sulfate) (PEDOT_SDS) modified electrode was prepared by electrochemical deposition method. Hereafter, the above-modified electrode denoted as PEDOT_SDS–nanoAg electrode. The nanoAg formation was confirmed by UV–vis spectroscopy and it shows surface plasmon resonance (SPR) peak at 375 nm. The surface morphology, film thickness, and surface roughness of PEDOT_SDS–nanoAg electrode were studied using atomic force microscopy (AFM). The PEDOT_SDS–nanoAg electrode has been utilized as a platform to immobilize electrochemically active mediator, Meldola Blue (MDB), by means of electrostatic trapping and the electrode denoted as PEDOT_SDS–nanoAg–MDB electrode. At 750 s of silver deposition time, reversible redox peak of MDB, which existed on PEDOT_SDS–nanoAg–MDB electrode, was obtained and that time was found to be an optimum. The electrochemical properties of the PEDOT_SDS–nanoAg–MDB electrode were studied by cyclic voltammetry (CV). The PEDOT_SDS–nanoAg–MDB electrode was investigated for catalytic application, which shows electrocatalytic activity towards the oxidation of NADH (dihydronicotinamide adenine dinucleotide) with 650 mV decrease in overpotential in comparison with bare glassy carbon electrode. Using the PEDOT_SDS–nanoAg–MDB electrode, the amperometric measurements were performed at an applied potential of −0.05 V and a linear response of NADH was obtained in the range from 10 to 560 μM with a limit of detection (LOD) of 0.1 μM. The PEDOT_SDS–nanoAg–MDB electrode showed diminished response to its interferences.