The Zika virus (ZIKV) is a global health threat due to its rapid spread and severe health implications, including congenital abnormalities and neurological complications. Differentiating ZIKV from other arboviruses such as dengue virus (DENV) is crucial for effective diagnosis and treatment. This study presents the development of a biosensor for detecting the ZIKV non-structural protein 1 (NS1) using gold nanoparticles (AuNPs) functionalized with monoclonal antibodies employing dynamic light scattering (DLS). The biosensor named ZINS1-mAb-AuNP exhibited specific binding to the ZIKV NS1 protein, demonstrating high colloidal stability indicated by a hydrodynamic diameter (DH) of 140 nm, detectable via DLS. In the absence of the protein, the high ionic strength medium caused particle aggregation. This detection method showed good sensitivity and specificity, with a limit of detection (LOD) of 0.96 µg mL-1, and avoided cross-reactivity with DENV2 NS1 and SARS-CoV-2 spike proteins. The ZINS1-mAb-AuNP biosensor represents a promising tool for the early and accurate detection of ZIKV, facilitating diagnostic and treatment capabilities for arboviral infections.