Split G‑Quadruplex\nProgrammed Recyclable AIE-Biosensor\nfor Label-Free Detection of miRNA in Acute Kidney Injury

We herein rationally designed a target-recyclable AIE-biosensor\nbased on a split G-quadruplex for label-free detection of miRNA in\nacute kidney injury. Initially, the PG was in an “OFF”\nstate, and the two split segments (G4-a and G4-b) of G4 were tethered\nat the two terminals of P1 and far away from each other due to the\nrigid duplex structure formed by the partially complementary intermediate\nsequences of P2 and P1, bringing MG with quenched fluorescence. In\nthe presence of target, the 5′-PO<sub>4</sub> P2 was displaced\nfrom PG probe and competitively hybridized with target, which led\nto G4-a and G4-b tending to form an intact intermolecular G-quadruplex,\nproviding sites for MG intercalation, thus generating an activated\n“ON” fluorescence signal due to the restriction of intermolecular\nmotion. Successively, relying on the λ-exonuclease (λ-Exo)\ncleavage reaction-assisted target recycling, more amounts of targets\nwill be liberated, accompanied by forming more G-quadruplex and binding\nmore MG, resulting in a strong fluorescence signal, further realizing\nthe sensitive detection of the targets. As a proof of concept, miRNA-21\nwas chosen as the model target. Endowing with the precise target recognition\nand efficient cleavage activity of λ-Exo, the AIE-biosensor\nexhibited excellent detection sensitivity and specificity, which could\nquantitatively detect miRNA-21 down to 10.36 fM with a single mismatch\nspecificity. The results revealed that the distinctive attributes\nof noninvasive, simple, and efficient in this G-quadruplex-based AIE-biosensor\noffered promising prospects for extensive applications in AKI screening\nand early clinical diagnosis.