Peptide Microarray-Based Metal Enhanced Fluorescence Assay for Multiple Profiling of Matrix Metalloproteinases Activities.

Matrix metalloproteinases (MMPs) are closely associated with cancer cell invasion and metastasis. Herein, a fluorescence resonance energy transfer (FRET)-peptide microarray-based metal enhanced fluorescence (MEF) assay is proposed for multiple and sensitive profiling of MMPs activities on a novel Au/Ag@SiO2 substrate. The Au/Ag@SiO2 substrate is prepared by electroless deposition of silver on gold nanoparticle (GNP) seeds, followed by SiO2 shell coating and surface functionalization. The specific FRET peptides are spotted on the Au/Ag@SiO2 substrate to sensitively detect MMPs (MMP-2, -3, -7, -9, -14) via fluorescence recovery by the MMP cleavage of quenched peptide motifs and further enhanced by MEF. Under the optimal conditions, the limits of detection are 12.2 fg mL-1 for MMP-2, 60 pg mL-1 for MMP-3, 0.22 pg mL-1 for MMP-7, 102 fg mL-1 for MMP-9, and 0.68 ng mL-1 for MMP-14, respectively. The practicability of the FRET-peptide microarray-based MEF assay is demonstrated by profiling of multiplexed MMPs activities in various cell lines and clinical thyroid tissue samples of papillary thyroid carcinoma (PTC) patients and thyroid nodules (TN) patients, and satisfactory results are obtained.