Tetrahedral DNA-Enhanced Interparticle Rolling Machine for High-Efficiency Human Neutrophil Elastase SERS Ratiometric Sensing in Serum and Tissue

The development of DNA rolling machines with high rolling efficiency for ratiometric biosensing is of great significance for the accurate diagnosis and evaluation of diseases. Herein, an interparticle DNA rolling machine constructed by well-oriented and ordered DNA nanorollers guided by tetrahedral DNA was exploited for high-efficiency lung tumor-related human neutrophil elastase (HNE) SERS ratiometric sensing. In this design, tetrahedral DNA with blocked DNAzyme was assembled on AuNPs to engineer well-oriented and ordered walking DNA nanorollers (WDNs) endowed with high collision efficiency and accessibility, significantly improving the reaction kinetics and rolling efficiency. When the DNAzyme "leg DNA" on WDNs were activated through the multivalent DNA converted by target HNE, the activated WDNs with polyoriented walkers rolled efficiently along directional hairpin "track DNA" on magnetic NPs (H1@MNPs) that served as continuously "cleaving-rolling-assembly" specific substrates (CRAS), constructing an interparticle rolling machine and finally converting HNE into a ratiometric SERS signal in a nanogap-rich silver nanoisland substrate. The CRAS increased the reaction direction and local concentration, improving the accessibility and signal. The interparticle relative motion with nonplanar polyoriented walking arms weakens the derailment of rolling "leg DNA", improves the processivity, and amplifies capability. Moreover, nanogap-rich silver nanoisland SERS substrates promote the formation of high-density hot spot domains, further improving the detection sensitivity. Of note, the rolling machine ratiometric biosensor successfully measures the HNE with a detection limit of 0.25 pM and can screen inhibitors and discriminate the HNE levels in serum and tissue of normal and lung tumor patients, suggesting that the biosensor provides an effective tool for early diagnosis, prognostic evaluation, and drug discovery of lung tumor.