Enzyme channeling assays

Receptors or enzymes are the preferred dmg targets since binding and enzyme inhibitor assays are standard in a research laboratory, they can be established easily and offer the possibility of automation. Transport proteins, ion channels and transcription factors are less attractive targets for drug screening programs, because automation is costly and not always possible. In addition, the requirement for highly sophisticated equipment and expertise is decisive. 

I Gibbons, R Armenta, RK DiNello, et al. Nonseparation enzyme channeling immu-nometric assays. Meth Enzymol 136 93, 1987. 

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... 

Wright, J.D., Rawson, K.M., Ho, W.O., Athey, D., and McNeil, C.J., 1995. Specific binding assay for biotin based on enzyme channelling with direct electron transfer electrochemical detection using horseradish peroxidase. Biosensors and Bioelectronics. 10 495-500. 

Under basal conditions, PKC is predominantly a cytoplasmic protein. Upon activation by Ca2+ or DAG, the enzyme associates with the plasma membrane, the site of many of its known physiological substrates, including receptors and ion channels. In fact, the translocation of PKC from the cytoplasm to the membrane has long been used as an experimental measure of enzyme activation. Such translocation has often been assayed by phorbol ester binding phorbol esters are tumor-promoting agents that selectively bind to and activate PKC. The molecular basis of the translocation of PKC from the cytoplasm to the plasma membrane has been solved. Subsequent to activation, PKC binds with high affinity to a series of membrane-associated proteins, termed receptors for... 

Affinities/functional activities at relevant molecular targets Pharmacological profiling — binding or functional assays at receptors, enzymes, ion channels, transporters Wakefield et al.,-83 Bowes et al.20... 

Receptor ligand and ion channel binding, enzyme assays, etc. 

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