A Peptide Cleavage-Based Ultrasensitive Electrochemical Biosensor with an Ingenious Two-Stage DNA Template for Highly Efficient DNA Exponential Amplification.

The direct transduction of a peptide cleavage event into DNA detection has always produced output DNA with some amino acid residues, which influence the DNA amplification efficiency in view of their steric hindrance effect. Here an ingenious two-stage DNA template was designed to achieve highly efficient DNA amplification by utilizing the DNA exponential amplification reaction (EXPAR) as a model. The usage of a two-stage DNA template not only accomplished the traditionally inefficient EXPAR triggered by output DNA with some amino acid residues but also simultaneously produced a newly identical DNA trigger without any amino acid residues to induce an extra efficient EXPAR, which significantly improved the DNA amplification efficiency, realizing the ultrasensitive detection of the target. On the basis of the proposed highly efficient DNA amplification strategy, a novel peptide cleavage-based electrochemical biosensor was constructed to ultrasensitively detect matrix metalloproteinases-7 (MMP-7). As a result, this developed assay demonstrated excellent sensitivity with a linear range from 0.1 pg·mL-1 to 50 ng·mL-1 and a detection limit down to 0.02 pg·mL-1, which paved a novel avenue for constructing ultrasensitive peptide cleavage-based biosensors.