Multiple electrochemical detection

Multiple electrodes have been used to obtain selectivity in electrochemical detection. An early example involved the separation of catecholamines from human plasma using a Vydac (The Separation Group Hesperia, CA) SCX cation exchange column eluted with phosphate-EDTA.61 A sensor array using metal oxide-modified surfaces was used with flow injection to analyze multicomponent mixtures of amino acids and sugars.62 An example of the selectivity provided by a multi-electrode system is shown in Figure 2.63... 

B. Munge, G. Liu, G. Collins, and J. Wang, Multiple enzyme layers on carbon nanotubes for electrochemical detection down to 80 DNA copies. Anal. Chem. 77, 4662-4666 (2005). 

The unique practical properties of adsorption have promoted its extensive use in genetic analysis. The disadvantages of adsorption with respect to covalent immobihzation are mainly that (1) nucleic acids may be readily desorbed from the substrate, and (2) base moieties may be unavailable for hybridization if they are bonded to the substrate in multiple sites [34]. However, the electrochemical detection strategy based on the intrinsic oxidation of DNA requires the DNA to be adsorbed in close contact with the electrochemical substrate by multi-point attachment. This multi-site attachment of DNA can be thus detrimental for its hybridization but is crucial for the detection based on its oxidation signals. 

Moreover, the unique adsorption properties of GEC allowed the very sensitive electrochemical detection of DNA based on its intrinsic oxidation signal that was shown to be strongly dependent of the multi-site attachment of DNA and the proximity of G residues to GEC [100]. The thick layer of DNA adsorbed on GEC was more accessible for hybridization than those in nylon membranes obtained with genosensors based on nylon/GEC with a changeable membrane [99,101,102]. Allhough GEC has a rough surface, it is impermeable, while nylon is more porous and permeable. DNA assays made on an impermeable support are less complex from a theoretical standpoint [7] the kinetics of the interactions are not compUcated by the diffusion of solvent and solutes into and out of pores or by multiple interactions that can occur once the DNA has entered a pore. This explained the lower hybridization time, the low nonspecific adsorplion and the low quantity of DNA adsorbed onto GEC compared to nylon membranes. 

MesoScale Discovery (MSD) succeeded in introducing product with a similar technology approach based upon ruthenium redox-mediated electrochemical detection (Figure 2.14). MSD is a joint venture of its parent company, MesoScale, and IGEN, a company that pioneered much of fhe work on electrochemical detechon based on the ruthenium redox system. MSD s Multi-Spot plates contain antibodies immobilized on multiple working electrode pads within each well, allowing each spot within the well to serve as an individual assay. Multiplexed cytokine immxmoassays can be performed in 96-well (4,7, or 10 spots per well) patterns with detection limits of 1 to 10 pg/mL and a linear dynamic range up to 3,000 pg/mL. Both 24-and 384-well electrode systems are available. 

Electrochemical detectors for liquid chromatography have reached a level of maturity in that thousands of these devices are used routinely for a variety of mundane purposes. Nevertheless, the technology is advancing rapidly in several respects. Multiple electrode and voltammetric detectors have been developed for more specialized applications. Small-volume transducers based on carbon fiber electrodes are being explored for capillary and micropacked columns. Recently, electrochemical detection has also been coupled to capillary electrophoresis [47]. Finally, new electrode materials with unique properties are likely to afford improved sensitivity and selectivity for important applications. 

Ikarashi et al. reported the development of a liquid chromatography multiple electrochemical detector (LCMC) and its application in neuroscience [197]. The system developed consisted of four parallel liquid chromatographic systems equipped with multiple electrochemical detectors, and the fourth was used for the assay of acetylcholine with a detection limit of 0.1-0.4 pmol. 

W.A. Bartlett, Effects of mobile phase composition on the chromatographic and electrochemical behaviour of catecholamines and selected metabolites reversed-phase ion-paired high-performance liquid chromatography using multiple-electrode detection, J. Chromatogr, 493, 1-14 (1989). 

Electrochemical detectors were reported used by 21% of the respondents to the detector survey (47). Electron transfer processes offer highly sensitive and selective methods for detection of solutes. Various techniques have been devised for this measurement process, with the most popular being based on the application of a fixed potential to a solid electrode. Potential pulse techniques, scanning techniques, and multiple electrode techniques have all been employed and can offer certain advantages. Two excellent reviews of electrochemical detection in flowing streams have appeared (59,60), as well as a comprehensive chapter in a series on liquid chromatography (61). 

Swanek, F.D., Chen, GY., and Ewing, A.G, Identification of multiple compartments of dopamine in a single ceU by CE with scanning electrochemical detection. Anal. Chem., 68, 3912, 1996. 

Recently, electrochemical detection methods, namely, conductimetry, amperometry, and potentio-metry, have also become accessible. All three variants of electrochemical detection are intrinsically simpler than the optical methods, and their success depends highly on the electrode materials and designs used. Conductivity detection relies on measurement of the differences between the conductivities of the analyte and the separation electrolyte this provides a direct relationship between migration times and response factor, and makes this detector universal. On the contrary, amperometric detection is restricted to electroactive species and potentiometric detection is not possible for certain small ions with multiple charges. Conductimetric detection works better for inorganic compounds since the higher mobility of... 

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