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Exosomes carrying abundant information have aroused great interest as effective biomarkers in liquid biopsy and are therefore ideal candidates for the early diagnosis of cancer and treatment monitoring. Herein, we developed a sensitive electrochemical biosensor using in situ generation of Fe₄[Fe(CN)6]₃ (Prussian Blue) on the surface of Ti3C2 MXene (two-dimensional transition-metal carbides) as hybrid nanoprobes (PB-MXene) for the detection of exosomes and their surface protein. A CD63 aptamer-modified poly(amidoamine) (PAMAM)-Au NP electrode interface was fabricated that can specifically bind with CD63 protein on the exosomes derived from OVCAR cells. In addition, the CD63-modified Ti3C2 MXene was used as a nanocarrier to accommodate numerous aptamers and was adsorbed on the exosomes. The Ti3C2 MXene can realize the in situ generation and high-efficiency loading of PB and further amplify the electrochemical signal at a low potential, thus avoiding the interference of the electrochemical active species. The dual amplification effect enables highly selective and sensitive electrochemical detection of exosomes. The limit of detection (LOD) was 229 particles μL-1 with a linear range from 5 × 102 particles μL-1 to 5 × 105 particles μL-1. An electrochemical biosensor can detect exosomes secreted by various cancer cells such as HeLa, OVCAR and BT474, and shows a high specificity even in serum samples, thus demonstrating its great potential in the application of clinical diagnostics. This proposed electrochemical biosensor provides a facile and efficient tool for the early diagnosis of cancers.