Enzyme Assay in Microfluidics

Enzyme Assay in Microfluidics, Fig. 3 Schematic of operations inside the microdroplet platform, (a) The mixing of two reagents (amino acid mix and nucleic acid mix) while the droplets are generated, (b) Droplet fusion between OpdA-containing droplets and coiunaphos... 

Enzyme Assay in Microfluidics, Fig. 4 (In black) Fluorescence measurement of the microdroplet inside the microfluidic chip befOTe (a) and after (b) droplet merging with the coumtqthos solution. The enzymatic reaction produced lluoresctait chlraferon inside some of the dn lets. 

Xue, Q., Wainright, A., Gangakhedkar, S., Gibbons, I., Multiplexed enzyme assays in capillary electrophoretic single-use microfluidic devices. Electrophoresis 2001, 22(18), 4000-4007. 

Wu N, Courtois F, Surjadi R, Oakeshott J, Peat TS, Easton CJ, Abell C, Zhu Y (2011) Enzyme synthesis and activity assay in microfluidic droplets on a chip. Eng Life Sci 11 157-164... 

The first type of enzyme application in microfluidics is chemical sensing. Sensors can be constmeted in cases where an enzyme turns over a particular smaU-molecule substrate to produce a product quantifiable by fluorescence, chemiluminescence, absorbance spectroscopy, or electrochemical detectors. In cases where the substrate is not detectable itself, an enzymatic product can often be coupled to another enzyme that produces a detectable product. For example, there are a wide variety of small molecules (such as nutrients, amino acids, and sugars) that can be coupled to the chemiluminescent reaction of luminol and peroxide in the presence of horseradish peroxidase. These enzyme-substrate assays were the first to be adapted to microfluidic devices — a great number of small-molecule sensors have been developed based on microfluidic channels with electroosmotic or hydrodynamic flow, and pre-loaded microfluidic cartridges containing nanoliter volumes of reagents have... 

Apart from immunoassays, enzyme assays can also be used to detect certain substrates in a clinical diagnostic setting. The benefits of performing enzymatic assays on microchips are the analytical power and minimal reagent use in microfluidic systems combined with the selectivity and amplification factors that come with biocatalysis. 

Schilling, E.A., Kamholz, A.E., Yager, P., Cell lysis and protein extraction in a microfluidic device with detection by a fluorogenic enzyme assay. Anal. Chem. 2002, 74(8), 1798-1804. 

In addition to commercially production, a great deal of research and development work on biochips has been going on both in industry and in academia. Genomic analysis of DNA and RNA continues to be the focus of interest, but more and more effort is being spent on proteomic analysis of proteins and peptides. Several enzyme assays and immunoassays designed based on microarray-based systems with simple microfluidic control are close to commercialization. They can be a vital tool in clinic diagnostics, drug discovery, and biomedical research. 

Lin, S. S., Fischl, A. S., Bi, X. H., and Parce, W., Separation of phospholipids in microfluidic chip device apphcation to high-throughput screening assays for lipid-modifying enzymes. Analytical Biochemistry 314, 97-107, 2003. 

Enzyme assays Enzyme synthesis In vitro com-partmentalization Microfluidics Water-in-oil emulsion... 

Quantitation of small molecules with enzymatic methods provides insight into the ccMicentration and activity of the proteins associated with those molecules. A great variety of these enzymatic assays have been carried out in microfluidic devices. Another function of enzymatic assays is in kinetics measurements of properties of enzymes such as the Michaelis-Menten constant (the concentration of substrate when the reactimi rate is half the maximum rate) and the turnover number (the number of moles of substrate that are converted to product per catalytic site per unit time) are vital to understanding the mechanics of the proteome and are used to characterize the effects of known drugs and discover new ones. 

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