The integration of hybridization chain reaction (HCR) with fluorogenic silver nanoclusters (AgNCs) in an aggregation induced emission (AIE)-based nanosensor for sex determination and its forensic application

We have designed a novel turn-on label-free nanosensor for sex determination by targeting DYS14 marker found on Y chromosome via integrating hybridization chain reaction (HCR) and silver nanoclusters (AgNCs). AgNCs are used as fluorophores, and for the first time, they are formed separately on a DNA template sequence other than the hairpins of HCR, to fulfill their sensitive synthesis and drive their aggregation induced emission (AIE) enhancement. While HCR acts as a simple, enzyme-free and isothermal amplification technique. The working principle is based on the hybridization of the target, when present, with a part of the hairpin H1 causing its opening, and then the other fragments open another hairpin H2 to hybridize with. As a last step, the free fragment found on H2 hybridizes with that attached to the nucleation sequence of AgNCs on the free DNA probes. As a result, a double stranded HCR product is formed, and the nanoclusters are placed in specific positions in a close proximity on it. This accumulation causes the phenomenon of AIE, and finally, the fluorescence emission intensity of AgNCs increases. This designed bioassay has the advantages of simple operation, cost effectiveness, rapid results, and good stability and specificity. Furthermore, it is the first time; an optical biosensor is introduced for this gene. Moreover, the detection limit of this assay is as low as 0.87 pM, and it also showed satisfying performance in real samples. This platform holds great promise for the application of label-free HCR/AgNCs nanosensors to other biomarkers.