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A novel electrochemical nano\u2010biosensor was developed for PKU detection, based on signal amplification using nanomaterials, e.g. gold nanoparticles (AuNPs) decorated on the reduced graphene oxide sheet on the screen\u2010printed carbon electrode. The fabrication steps were checked by field emission scanning electron microscope imaging as well as cyclic voltammetry analysis. The specific alkanethiol single\u2010stranded DNA probes were attached by self\u2010assembly methodology on the AuNPs surface and Oracet blue was used as an intercalating electrochemical label. The results showed the detection limit of 21.3 fM and the dynamic range of 80\u20131200 fM. Moreover, the selectivity results represented a great specificity of the nano\u2010biosensor for its specific target DNA oligo versus other non\u2010specific sequences. 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