|Title||Design optimization for bioMEMS studies of enzyme-controlled metabolic pathways.|
|Publication Type||Journal Article|
|Year of Publication||2008|
|Authors||Luo, X, Berlin, DLarios, Buckhout-White, S, Bentley, WE, Payne, GF, Ghodssi, R, Rubloff, GW|
|Date Published||2008 Dec|
|Keywords||Chitosan, Enzymes, Immobilized, Escherichia coli, Microfluidic Analytical Techniques, N-Glycosyl Hydrolases|
Biological microelectromechanical systems (bioMEMS) provide an attractive approach to understanding and modifying enzymatic pathways by separating and interrogating individual reaction steps at localized sites in a microfluidic network. We have previously shown that electrodeposited chitosan enables immobilization of an enzyme at a specific site while maintaining its catalytic activity. While promising as a methodology to replicate metabolic pathways and search for inhibitors as drug candidates, these investigations also revealed unintended (or parasitic) effects, including products generated by the enzyme either (1) in the homogeneous phase (in the liquid), or (2) nonspecifically bound to microchannel surfaces. Here we report on bioMEMS designs which significantly suppress these parasitic effects. To reduce homogeneous reactions we have developed a new packaging and assembly strategy which eliminates fluid reservoirs that are commonly used for fluidic interconnects with external tubing. To suppress reactions by nonspecifically bound enzyme on microchannel walls we have implemented a cross-flow microfluidic network design so that enzyme flow for assembly and substrate/product for reaction share only the region where the enzyme is immobilized at the intended reaction site. Our results show that the signal-to-background ratio of sequential enzymatic reactions increases from 0.72 to 1.28 by eliminating the packaging reservoirs, and increases to 2.43 by separating the flow direction of enzymatic reaction from that of enzyme assembly step. These techniques can be easily applied to versatile microfluidic devices to minimize parasitic reactions in sequential biochemical reactions.
|Alternate Journal||Biomed Microdevices|