Abstract

Thrombin has been implicated in atherosclerotic disease development. However, thrombin activity detection is currently limited because of the lack of convenient fluorescent probes. We developed a label-free fluorescent method to assay thrombin activity on the basis of a designed peptide probe with a thrombin-cleavable peptide sequence and a cysteine terminus. The peptide probe can be conjugated to DNA-templated silver nanoclusters (DNA-AgNCs) through Ag-S bonding; as a result, the fluorescence of DNA-AgNCs was enhanced. As the DNA-AgNCs-peptide conjugate was adsorbed to graphene oxide (GO), the enhanced fluorescence of DNA-AgNCs was quenched. Once the peptide probe was cleaved by thrombin, the resulting release of the DNA-AgNCs from the surface of GO restored the enhanced fluorescence. Thrombin can be determined with a linear range of 0.0-50.0 nM with a detection limit of 1 nM. The thrombin-sensitive probe with a cysteine terminus may be developed into probes to detect other proteases.

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