Measurement of Enzyme Kinetics Using a Continuous-Flow Microfluidic System
- Title
- Measurement of Enzyme Kinetics Using a Continuous-Flow Microfluidic System
- Author
- 성기훈
- Issue Date
- 2003-07
- Publisher
- AMER CHEMICAL SOC
- Citation
- ANALYTICAL CHEMISTRY, v. 75, issue. 13, page. 3161-3167
- Abstract
- This paper describes a microanalytical method for determining enzyme kinetics using a continuous-flow microfluidic system. The analysis is carried out by immobilizing the enzyme on microbeads, packing the microbeads into a chip-based microreactor (volume similar to 1.0 nL), and flowing the substrate over the packed bed. Data were analyzed using the Lilly-Hornby equation and compared to values obtained from conventional measurements based on the Michaelis-Menten equation. The two different enzyme-catalyzed reactions studied were chosen so that the substrate would be nonfluorescent and the product fluorescent. The first reaction involved the horseradish peroxidase-catalyzed reaction between hydrogen peroxide and N-acetyl-3,7-dihydroxyphenoxazine (amplex red) to yield fluorescent resorufin, and the second the beta-galactosidase-catalyzed reaction of nonfluorescent resorufin-beta-D-galactopyranoside to yield D-galactose and fluorescent resorufin. In both cases. the microfluidics-based method yielded the same result obtained from the standard Michaelis-Menten treatment. The continuous-flow method required similar to10 muL of substrate solution and 10(9) enzyme molecules. This approach provides a new means for rapid determination of enzyme kinetics in microfluidic systems, which may be useful for clinical diagnostics, and drug discovery and screening.
- URI
- https://pubs.acs.org/doi/10.1021/ac034155bhttps://repository.hanyang.ac.kr/handle/20.500.11754/156124
- ISSN
- 0003-2700
- DOI
- 10.1021/ac034155b
- Appears in Collections:
- COLLEGE OF ENGINEERING SCIENCES[E](공학대학) > BIONANO ENGINEERING(생명나노공학과) > Articles
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