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Biopolymer films have drawn growing demand for their application in the point of care domain owing to their biocompatibility, eco-friendly, and eligibility for in vivo analyses. However, their poor conductivity restricts their sensitivity in diagnostics. For high-quality electrochemical biosensor monitoring, two vital factors to be greatly paid attention are the effective merge of amplification modifiers with transducing surface and the superior linking across the recognition interface. Here, we introduce an enzyme-free electrochemical biosensor based on electrosynthesized biocompatible WO3/poly glutamic acid nano-biocomposites to address the hardships specific to the analysis of circulating proteins clinical samples. In addition to its green synthesis route, the poor tendency of both components of the prepared nano-biocomposite to amine groups makes it excellent working in untreated biological samples with high contents of proteins. Several electrochemical and morphological investigations (SEM, EDX, and dot mapping) were fulfilled to gain a reliable and trustful standpoint of the framework. By using this nanobiosensor, the concentration of HER-2 was detectable as low as 1 fg mL−1 with a wide linear response between 1 ng mL−1 and 1 fg mL−1. Meanwhile, the protocol depicted ideal specificity, stability, and reproducibility for the detection of HER-2 protein in untreated serum samples of breast cancer patients. [ABSTRACT FROM AUTHOR]