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Abstract Circulating miRNAs in the blood can regulate disease development and thus indicate disease states via their various expression levels. For these reasons, circulating miRNAs constitute useful biomarkers, and an approach to the accurate detection of circulating miRNAs is attractive in the diagnosis and treatment of diseases. However, methods for clinical detecting of circulating miRNA that take both sensitivity and practicality into account are still needed. Therefore, we aimed herein to solve some inherent problems in the actual detection using a robust surface-enhanced Raman scattering (SERS) platform with integrated nucleic acid amplification and nanoparticle aggregation to construct 3D hotspots for improving performance of analyzing circulating miRNAs. After target recognition and initial signal amplification by DNAzyme, we observed that release triggered an open hairpin DNA on gold nanoparticles (AuNPs), which then promote AuNP aggregation, causing the accumulation of a large number of hotspots in three-dimention. The SERS biosensor achieved a better performance than the sandwich-type separation detection, with a low detection limit (0.37 fM) and a broad linear range (1 fM–10 nM) in liquids. This SERS platform can be used as a powerful tool for the detection of circulating miRNAs, and it can be used to improve the sensitivity and accuracy of various clinical-disease diagnoses. Graphical Abstract