Homogeneous Immunoassay for Detection of TNT and Its Analogues on a Microfabricated Capillary Electrophoresis Chip

Abstract

A homogeneous immunoassay for TNT and its analogues is developed using a microfabricated capillary electrophoresis chip. The assay is based on the rapid electrophoretic separation of an equilibrated mixture of an anti-TNT antibody, fluorescein-labeled TNT, and unlabeled TNT or its analogue. The band intensities of the free fluorescein-labeled TNT and of the antibody-antigen complex reveal the relative equilibrated concentrations. Titration of the anti-TNT antibody with a fluorescein-labeled TNT derivative yields a binding constant of (3.9 +/- 1.3) x 10(9) M(-1). The dissociation rate constant of the complex is determined by kinetic capillary electrophoresis using a folded channel and a rotary scanner to interrogate the separation at multiple time points. The dissociation rate constant is found to be 0.035 +/- 0.005 s(-1), and the resulting binding rate constant is (1.4 +/- 0.7) x 10(7) M(-1) s(-1). Binding constants of TNT and five of its analogues are determined by competitive assays: TNT (4.3 +/- 2.6) x 10(8) M(-1); 1,3,5-trinitrobenzene (5.1 +/- 3.3) x 10(7) M(-1); picric acid (7.5 +/- 4.4) x 10(6) M(-1); 2,4-dinitrotoluene (7.9 +/- 4.0) x 10(6) M(-1); 1,3-dinitrobenzene (1.0 +/- 0.7) x 10(6) M(-1); and 2,4-dinitrophenol (5.1 +/- 3.0) x 10(4) M(-1). TNT and its analogues can be assayed with high sensitivity (LOD 1 ng/mL) and with a wide dynamic range (1-300 ng/mL) using this chip-based method.

Integration of a carbon microelectrode with a microfabricated palladium decoupler for use in microchip capillary electrophoresis/ electrochemistry

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Moment Analysis Theory for Size Exclusion Capillary Electrochromatography with Chemical Reaction of Intermolecular Interaction

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Electrochemical Sensor for Explosives Precursors’ Detection in Water

“…Spectroscopic approaches have been the most widely explored [2][3][4][5][6][7], but quartz crystal microbalance (QCM), ion chromatography [8], capillary electrophoresis [9], surface-enhanced Raman scattering (SERS) [10], nanotechnology based-methods (using molecularly imprinted polymers [11], nanotubes, or nanoparticles), and sensor techniques [12][13][14][15][16] have also been largely described. Amongst the different types of sensors developed to address this need, the electrochemical sensors [17][18][19] present the advantages of being fast, inexpensive, and adapted to miniaturization.…”