Catalytic hairpin assembly-mediated SERS biosensor for double detection of MiRNAs using gold nanoclusters-doped COF substrate

A novel surface-enhanced Raman scattering (SERS) and catalytic hairpin assembly (CHA) technique dependent dual-signal amplification method for double detection of microRNAs (miRNAs) was created in this work. When target miRNAs were present, CHA reactions could be triggered between the corresponding hairpin DNAs, fixing the SERS tags (two distinct Raman reporters, 4-mercaptobenzonitrile and 4-mercaptopyridine, which were prelabeled in the space between Au core and silver shell nanoparticles) onto the gold nanoclusters-doped covalent organic frameworks (AuNCs/COF) nanofiber substrate surface, which generated numerous "hot spots" and ultimately produced amplified SERS signals. Besides, with the help of the CHA cycles, much more target miRNAs were captured to improve the detection sensitivity and decrease the detection limit of multiplex detection down to 77 aM for miRNA-21 and 93 aM for miRNA-155, respectively. This SERS biosensor also exhibited a broad detection range (10-16 - 10- 12 M), good specificity, high reproducibility, as well as excellent recoveries in spiked human serum samples (in the range of 94.5-99.4%). The findings of our experiment suggested that the multiple signal amplification biosensors that were developed have significant potential for multiplex detection of cancer biomarkers (miRNAs and their analogs) in the fields of bioanalysis and clinical biomedicine.