An integrated magneto-fluorescent nanosensor for rapid and sensitive detection of tumor-derived exosomes

Cancer released-exosomes containing specific surface proteins have high value in tumor diagnosis and are considered a novel liquid biopsy biomarker. However, the development of rapid and sensitive quantitative methods for their determination remains challenging because of the low abundance of exosome surface protein. Herein, we developed a new platform using an integrated magneto-fluorescent exosome (iMFEX) nanosensor for rapid and sensitive detection of exosomes derived from cancers. First, magnetic beads were coated with a DNA tetrahedral lipid probe to efficiently capture exosomes. The rationally designed bifunctional aptamer specifically recognized exosomal PD-L1 protein and initiated catalytic hairpin assembly, thereby converting protein signals into H1/H2 duplexes. Finally, the H1/H2 duplexes activated the Cas12a protein trans-cleavage reporter FAM-TTATT-BHQ1 for fluorescence signal amplification. By integrating magnetic separation and fluorescence signal amplification, the proposed iMFEX nanosensor exhibited excellent specificity and sensitivity toward PD-L1 positive exosomes, ranging from 2.86 × 103 to 2.86 × 107 particles/μL, with a detection limit of 1.71 × 103 particles/μL. Using the iMFEX nanosensor, we were able to effectively track the dynamic changes in exosomal PD-L1 expression induced by reagents and to distinguish patients with non-small cell lung cancer from healthy individuals, thus demonstrating the platform’s potential for clinical application.