DNAzyme-Based Microscale Thermophoresis Sensor

Microscale thermophoresis (MST) technology has emerged as a powerful growing method in a molecular interaction study by measuring fluorescence responses of molecules inside a capillary to infrared (IR) laser heating with the benefits of rapid ratiometric measurement, separation-free, no immobilization, and low sample consumption. Combining the advantages of RNA cleaving DNAzymes in target recognition and enzymatic catalysis and the strength of MST technology for fluorescence signaling, here, we reported a DNAzyme-based MST method for sensitive target detection. We introduced a fluorescein terminal label at the RNA-cleaving DNAzyme, and the substrate was linked to DNAzyme together with a poly-T sequence in a unimolecular design or not conjugated with DNAzyme in a bimolecular design. The presence of the cofactor activated DNAzyme to catalytically cleave the substrate, causing molecular structure alteration and significant changes in MST signals. This DNAzyme MST sensor enabled sensitively detecting activator targets Pb2+ and L-histidine, with a detection limit of 49 pM Pb2+ and 3.9 mu M L-histidine. This biosensing strategy is universal and promising for wide applications.

Automated summary

Microscale thermophoresis (MST) technology has been used for studying molecular interactions through fluorescence responses of molecules to infrared laser heating. In this study, a DNAzyme-based MST method was developed for sensitive target detection, by combining the advantages of RNA cleaving DNAzymes and MST technology. This biosensing strategy showed high sensitivity and has broad applications.