Tandem Hybridization Chain Reaction and Selective Coordination Enable Fluorescence Detection of Exosomes in Lung Cancer

Liquid biopsy based on tumor-derived exosomes (TDEs) is potentially effective in assisting early cancer diagnosis. However, accurate quantification of the approach remains challenging due to the low concentration and heterogeneity of TDEs. This work involved designing a homogeneous enzyme-free fluorescent sensing strategy by targeting programmed cell death ligand 1 (PD-L1) that was overexpressed on the surface of TDEs, coupled with the tandem hybridization chain reaction (HCR) and the selective coordination of Cu2+. The aptamer was used as a specific recognition probe for binding with the target protein, thereby regulating the tandem HCR reactions and achieving signal amplification. The coordination between Cu2+ and double-stranded DNA (dsDNA) can further regulate the fluorescence intensity of the signal molecule, calcein, for precise quantification of TDEs. The results showed that the detection limit (LOD) for TDEs can be as low as 100 particles/mL. Moreover, the proposed strategy can accurately differentiate between lung cancer patients (n = 25) and healthy donors (n = 12), with a sensitivity of 92%, specificity of 100%, and an area under the receiver operating characteristic curve (AUC) of 0.963. The fluorescence analysis also demonstrated a strong correlation with the computed tomography scans and pathological analysis. Therefore, the proposed method in this study can provide more insights into assisting in the early diagnosis of lung cancer through TDEs-based liquid biopsy.