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Plasma deposited polyoxazoline thin films (POx) are a promising solution for the rapid, scalable, and substrate-independent immuno-functionalization of electrochemical biosensors. However, a major challenge in using POx thin films in electrochemical sensing is their inherently insulating nature. This work reports the incorporation of gold nanoparticles (AuNPs) between two layers of POx which enhances the conductivity of the films. The size of the AuNPs, their binding density on the POx underlayer, and the POx films’ intrinsic electrical resistance were all factors in improving the overall electrochemical response of the layered construction. Surface bound electrochemical measurements and conductive atomic force microscopy were conducted to uncover a possible mechanism for the observed nanoparticle-mediated electron transport through the insulating matrix. The primary contributor to increasing conductivity in …