A potent fluorescent biosensor integrating 3D DNA walker with localized catalytic hairpin assembly for highly sensitive and enzyme-free Zika virus detection

Zika virus (ZIKV) with worldwide distribution, high incidence and great harm, has emerged as a serious global health threat in the past few years. There is an urgent need for fast, low-cost and accurate diagnosis of ZIKV. Herein, a DNA walker integrated with localized catalytic hairpin assembly (LCHA) cascade amplification strategy is proposed as a new ZIKV assay platform satisfying the above requirements. In this strategy, the DNA walker is constructed by functionating AuNPs with locked walking strands and hairpin DNA track strands; LCHA is simply designed by fixed hairpin DNA probes H1 and H2 on a DNA tetrahedron. The target ZIKV RNA sequence initiates the DNA walker process, which generates numerous trigger sequences for activating the subsequent LCHA, generating an amplified fluorescence signal. This method allows the limit of ZIKV sequence detection down to 20 pM without enzymatic DNA amplification. Moreover, the assay can detect ZIKV from serum sample without RNA extraction, proving its practical application in biological analysis. • Integrated DNA walker with LCHA cascade amplification for ZIKV diagnostic. • Sensitive, fast and nucleic acids amplification-free method. • Successful development and validation with serum sample.