Gold nanoparticle enhanced fluorescence anisotropy for the assay of single nucleotide polymorphisms (SNPs) based on toehold-mediated strand-displacement reaction

► This is the first effort combining gold nanoparticle fluorescence anisotropy signal enhancement and the toehold-mediated strand-displacement reaction to develop a homogeneous assay for SNP detection. ► The drastic dynamic and kinetic difference in strand-displacement from toehold design was used to distinguish between the perfectly matched and the single-base mismatched strands. ► Free energy changes were calculated to elucidate the dependence of the differentiation ability on the mutation site in the toehold region. ► A solid negative signal change can be obtained for single-base mismatched strand in the dynamic range, and a more than 10-fold signal difference was observed in a mixed solution containing 100 times the mismatched strand.