Development of a matrix metalloproteinase-2 (MMP-2) biosensing system by integrating β-lactamase-mediated chromogenic reaction with common electronic components
- 주제(키워드) biosensor
- 발행기관 아주대학교
- 지도교수 윤현철
- 발행년도 2017
- 학위수여년월 2017. 2
- 학위명 석사
- 학과 및 전공 일반대학원 분자과학기술학과
- 실제URI http://www.dcollection.net/handler/ajou/000000024367
- 본문언어 영어
- 저작권 아주대학교 논문은 저작권에 의해 보호받습니다.
초록/요약
Matrix metalloproteinase-2 (MMP-2) plays an important role in extracellular matrix (ECM) degradation, which allows cancer cells to migrate out of the primary tumor to form metastases. Thus, it can serve as a biomarker for cancer diagnosis. Various methods have been developed to analyze MMP-2 activities, however, their wide applications for disease diagnosis have been hampered because high-end analytical equipment and labor-intensive processes are required. In this study, we developed an MMP-2 biosensing system by integrating an engineered autoinhibited β-lactamase-mediated chromogenic reaction into common electronic components such as a laser diode, solar cell, and multimeter. The autoinhibited β-lactamase was immobilized on a polymeric biosensing channel by a polydopamine coating and self-assembled monolayer methods. In the presence of MMP-2, the autoinhibited β-lactamase was turned to an active form which can hydrolyze CENTA, a chromogenic substrate of β-lactamase. As a result, the substrate color was changed from pale yellow (λmax = 340 nm) to dark yellow (λmax = 405 nm). By reading the interfered laser-light intensity, we were able to analyze MMP-2 activities precisely both with the samples prepared in a buffer solution and also those in urine. These results suggested that the developed system can be used for the quantitative analysis of enzyme activity related to cancer diagnosis.
more목차
Abstract ………………………………………………………………………… ⅰ
Contents ……………………………………………………………………… ⅱ
List of Figures ………………………………………………………………… ⅴ
Development of a matrix metalloproteinase-2 (MMP-2) biosensing system by integrating β-lactamase-mediated chromogenic reaction with common electronic components
1. Introduction
1.1 Matrix metalloproteinase-2 (MMP-2) …………………………… 1
1.2 Conventional MMP-2 assay …………………………………………2
1.3 Engineered autoinhibited β-lactamase …………………………… 3
1.4 Developed optical sensing system …………………………………5
1.5 Purpose of the research…………………………………………………7
2. Material and Methods
2.1 Materials and apparatus …………………………………………… 10
2.2 Construction of the optical transducer using common electronics components ………………………………………………………………… 11
2.3 β-lactamase expression and purification …………………………12
2.4 Fabrication of the reaction channel …………………………………13
2.5 Surface modification of the reaction channel ……………………14
3. Result and discussion
3.1 β-lactamase Activity Assay ……………………………………… 15
3.2 β-lactamase Immobilization on the Biosensing Channel Surface ……………………………………………………………………………20
3.3 Construction of an Optical Biosensing System for β-lactamase ……………………………………………………………………………23
3.4 Development of an MMP-2 Activity Analysis Method …………………………………………………………………………… 27
3.5 Analysis of Urinary MMP-2 Activity …………………………… 30
4. Conclusion ……………………………………………………………… 33
5. Reference ………………………………………………………………… 34
