Structure-switching aptamer triggering hybridization displacement reaction for label-free detection of exosomes

Exosomes play important roles in intercellular communications, tumor migration and invasion. However, the specific detection of cancer exosomes remains as a big challenge due to its low concentration in biofluids. Therefore, the sensitive and selective detection of cancer cells-derived exosomes has attracted growing attention owing to their potential in diagnostic and prognostic applications. Activatable strategies have received great attention for the detection of low abundant analytes due to their high sensitivity. Herein, based on molecular recognition between DNA aptamer and exosome surface biomarker (protein tyrosine kinase-7), a novel activatable and label-free strategy was designed for highly sensitive and specific sensing of exosomes. In this work, the target exosomes trigger strand replacement reaction to form G-quadruplex, which result in an obvious fluorescence enhancement of N-methylmesoporphyrin IX due to the bonding between G-quadruplex and N-methylmesoporphyrin IX. Under the optimum experimental conditions, the linear range for exosomes was measured to be 5.0 × 10⁵-5.0 × 10⁷ particles/μL and the detection limit (LOD) was calculated to be 3.4 × 10⁵ particles/μL (3σ). This assay possesses high specificity to distinguish exosomes derived from different cell lines, and has successfully been validated in patient and healthy plasma samples. Furthermore, the probe can effectively detect the exosomes in 30% fetal bovine serum, indicating that the biological matrix has a negligible effect on this method. This developed label-free, convenient and highly sensitive biosensor will offer a great opportunity for exosomes quantification in biological study and clinical application.