CRISPR/Cas12a-based biosensor for colorimetric detection of serum prostate-specific antigen by taking nonenzymatic and isothermal amplification

CRISPR/Cas12a-based biosensors have received intensive attention in the detection of nucleic acids, but their application for the detection of protein cancer biomarkers remains challenging. Here, we report an enzyme-free hybridization chain reaction (HCR)-powered CRISPR/Cas12a-based biosensor for colorimetric detection of serum prostate-specific antigen (PSA). The nonenzymatic and isothermal characteristics of HCR were utilized to transduce serum PSA into nucleic acids product, which was specifically recognized by the CRISPR/Cas12a system along with the activation of the trans-cleavage activity of Cas12a. Activated Cas12a cleaved the linker ssDNA between the gold nanoparticles (AuNPs)-DNA probes pair for inducing the dispersion of AuNPs with color change. The results showed that this assay was capable of the sensitive and selective detection of PSA in spiked samples and clinical samples in visual and quantitative modes, exhibiting a limit of detection (LOD) of 0.1 ng mL−1 (3δ/slope). This assay well integrates enzyme-free and isothermal amplification power of HCR, specific recognition and trans-cleavage activity of CRISPR/Cas12a system and distance-dependent optical properties of AuNPs into a single sensing system, serving as a solid basis for the development of nonenzymatic and isothermal amplified CRISPR/Cas system-based biosensor for the detection of protein cancer biomarkers.