Development of microbiochip for detection of metalloproteinase 7 using fluorescence resonance energy transfer

作者：Lee Seung Yong; Han Byoungwook; Park Chansoo; Jeong Je Sik; Ahn Jeong Jin; Ha Seung Mo; Hwang Seung Yong; Ahn Yoomin^*

来源：BioChip Journal, 2013, 7(2): 164-172.

DOI：10.1007/s13206-013-7210-z

• Microbiochip
• Matrix Metalloproteinase 7
• Fluorescence Resonance Energy Transfer (FRET)
• Quantum Dot (QD)
• Bead-based assay
• Enzyme assay

A protease is any enzyme that catalyzes the hydrolysis of proteins into smaller peptide fragments and amino acids, a process known as proteolysis. They are involved in a multitude of normal biological processes as well as in diseases, including cancer, stroke and infections. Here we present a microfluidic-based assay system to detect proteolytic activity using fluorescence resonance energy transfer (FRET) by quantum dot (QD)-peptide conjugates immobilized on microbeads. As an energy donor, QD was immobilized on the microbead surface by the avidin-biotin interaction. As an energy acceptor, the fluorophore-labeled peptide was then associated with QD, thus quenching the photoluminescence (PL) of the QD. The functionalized microbeads were introduced into the microbiochip and captured by a micropillar in the reaction chamber. In the presence of matrix metalloprotease-7 (MMP-7) as a model protease, the PL of QD quenched by fluorophore was recovered due to the proteolytic activity of MMP-7 in the fabricated microbiochip. Moreover, the FRET efficiency induced by MMP-7 was linearly dependent on the logarithmic concentration of MMP-7. This technology is not limited to sensing MMP-7, but could be used to monitor other protease activities (Schematic diagram).

• 出版日期2013-6-20