Ultraspecific and highly sensitive nucleic acid detection by integrating a DNA catalytic network with a label-free microcavity.

Nucleic acid detection with label-free biosensors circumvents costly fluorophore functionalization steps associated with conventional assays by utilizing transducers of impressive ultimate detection limits. Despite this technological prowess, molecular recognition at a surface limits the biosensors' sensitivity, specificity, and reusability. It is therefore imperative to integrate novel molecular approaches with existing label-free transducers to overcome those limitations. Here, we demonstrate this concept by integrating a DNA strand displacement circuit with a micron-scale whispering gallery mode (WGM) microsphere biosensor. The integrated biosensor exhibits at least 25-fold improved nucleic acid sensitivity, and sets a new record for label-free microcavity biosensors by detecting 80 pM (32 fmol) of a 22nt oligomer; this improvement results from the catalytic behavior of the circuit. Furthermore, the integrated sensor exhibits extremely high specificity; single nucleotide variants yield 40- to 100-fold lower signal. Finally, the same physical sensor was demonstrated to alternatingly detect 2 different nucleic acid sequences through 5 cycles of detection, showcasing both its reusability and its versatility.