Amplified Assembly of a Multifunctional 3D DNA Nanomachine for Electrochemical and Colorimetric Dual-Mode Assays

DNA nanomachines have been widely applied in biosensing; however, further enhancing their signal amplification capability and assay reliability remains a major challenge. Herein, a novel strategy was proposed to construct a multifunctional three-dimensional DNA nanomachine for the development of an electrochemical and colorimetric dual-mode biosensing method for detecting the estrogen pollutant 17β-estradiol. Aptamer recognition-triggered catalytic hairpin assembly, together with a cascade Nb.BbvCI-assisted DNA polymerization reaction, drove the amplified assembly of the DNA nanomachine composed of multiple triangular prism units. The abundant Mg²⁺-dependent DNAzymes formed on the nanomachine enabled its efficient walking on an iron-based metal-organic framework (Fe-MOF)-modified electrode, thereby generating an electrochemical signal. Meanwhile, numerous G-quadruplexes were formed accompanying the DNA nanomachine assembly, thereby enabling sensitive colorimetric signal readout through the excellent peroxidase-mimicking activity of the formed G-quadruplex/hemin DNAzymes, synergistically promoted by released Fe-MOFs. Consequently, the proposed method exhibited wide linear ranges of 0.01 pg mL^-1 to 1 ng mL^-1 (electrochemical) and 0.1 pg mL^-1 to 10 ng mL^-1 (colorimetric) with detection limits as low as 3.9 fg mL^-1 and 10.5 fg mL^-1, respectively. In addition, the method showed simple operation, excellent repeatability, and enhanced reliability through cross-validation between the dual detection modes. Therefore, this work provides a robust and sensitive approach for monitoring environmental estrogen pollutants.