A label-free fluorescent aptasensor based on HCR and G-quadruplex DNAzymes for the detection of prostate-specific antigen

Abstract

Prostate specific antigen (PSA) has been considered as the most potential serological biomarker for the early stage detection of prostate cancer. Here, a label-free fluorescence aptasensing strategy for detecting PSA based on hybridization chain reaction (HCR) and G-quadruplex DNAzymes has been developed. This designed strategy consists of three DNA probes, aptamer probe (AP), hairpin probe 1 (H1) and hairpin probe 2 (H2). In the presence of target PSA, the aptamer sequences in AP specifically recognized PSA to form a PSA–aptamer complex, causing an AP conformation change and thus releasing the initiator, which triggered the chain-like assembly of H1 and H2 that yielded extended nicked double-stranded DNA through HCR. Upon the addition of hemin, the G-rich segments at the end of H1 and H2 self-assembled into the peroxidase-mimicking hemin/G-quadruplex DNAzymes, which catalyzed the hydrogen peroxide-mediated oxidation of thiamine to give a fluorescence signal dependent on the concentration of PSA. Under optimal conditions, a limit of detection of 0.05 nM and a linear range from 0.1 nM to 1 nM (R² = 0.9942) were achieved by this assay. In addition, other interfering proteins, such as IgG, AFP and CEA, did not produce any significant change in the fluorescence intensity response, indicating good selectivity of this sensor for PSA detection. Finally, this proposed aptasensor was successfully used for diluted serum samples.