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In this work, based on the powerful cycle amplification cascades of proximity hybridization-induced hybridization chain reaction and catalyzed hairpin assembly, we engineered a nonenzymatic and ultrasensitive method which combined the Mg2+-DNAzyme recycling signal amplification for the analysis of the human prostate specific antigen. Herein, we adopted PSA-conjugates as triggers in the self-assembly process of two hairpin DNAs (H1, H2) into the products of the CHA which could activate the HCR to induce repeated hybridization. And both ends of each adjacent sequence of the HCR products could form a unit of Mg2+-DNAzyme which in presence of cofactor Mg2+ could recognize and cyclically cleave the hairpin probes in the solution and thus generate observably enhanced fluorescent signal. Benefit from the nucleic acid circuit amplification strategy, PSA of concentration low to 0.73 pg mL−1 was detected in this system. This homogeneous sensing method in solution avoid the use of the sophisticated equipment and complex operation, as well as addition of artificial enzyme, thus greatly reducing the constraints and complexity of experimental conditions. Moreover, considering most protein biomarkers in serum don't have their corresponding aptamers, this sensing method provide a general sensing approach for homogeneous sensitive detection of these important protein biomarkers which transfer rough antigen-antibody interactivity to smart signal amplification sensing strategies, thus exhibiting a remarkable prospect in clinical application.

An amplified fluorescence biosensing strategy for serum prostate specific antigen (PSA) was developed on the basis of DNAzyme. In presence of cofactor Zn2+, Zn2+ -dependent DNAzyme could cleave the hairpin substrate probes which were dispersed in solution and generate remarkable fluorescent signal. Taking advantage of the magnetic beads as a carrier, one target protein could bring plentiful hairpin substrate probes on to the electrode through a sandwich structure (Ab1/PSA/biotin-Ab2). Moreover, during the cleavage process of as formed DNAzyme, DNAzyme did not be destroyed and could further react with other hairpin probes, then generated continuous fluorescent signal. Benefited by this amplified strategy, the limit of detection (LOD) was low to 0.05 ng mL−1, which was much lower than our previous reports. This method could be applied to detect different protein biomarkers in serum without corresponding aptamers by changing the corresponding antibodies and thus showed a remarkable prospect in clinical application.