Peptide nucleic acid modulated fluorescence light-up DNA-Ag nanoclusters for sensitive and specific detection of DNA

Fluorescent silver nanoclusters, particularly DNA-templated silver nanoclusters (DNA-AgNCs), have garnered significant attention owing to their tunable fluorescence properties and excellent biocompatibility. Here we demonstrate that thiol-functionalized peptide nucleic acid (SH-PNA) with mixed base composition can significantly enhance the fluorescence emission intensity of C₁₂-AgNCs. This fluorescence enhancement induced by PNA can be effectively modulated through the hybridization of PNA with specific DNA sequences. Based on this finding, we have developed a fluorescence biosensor for sensitive and specific detection of DNA utilizing PNA-AgNCs probes, which enables highly efficient detection of single-base mutations in the TP53 gene fragment. Under optimal conditions, the fluorescence of the PNA-AgNCs probe exhibits a good linear relationship with the concentration of target DNA, achieving a detection limit of 1.3 nM and demonstrating inherent high specificity for single-base mutations. Regardless of the mismatch type, this method allows for the screening of mutant genes within 45 min. Furthermore, this detection strategy is operationally simple and has been successfully applied to the analysis of single-base mismatches in TP53 DNA from NCI-H661 lung cancer cells. In principle, by simply modifying the sequence of the PNA probe, this detection strategy can be readily extended to the detection of other nucleic acids.