Microcapillary reversed-displacement immunosensor for trace level detection of TNT in seawater

A reversed-displacement immunosensor to detect the explosive, 2,4,6-trinitrotoluene (TNT) using a chemically modified borosilicate glass microcapillary is described. The inner core of a glass microcapillary was modified with 3-aminopropyltriethoxysilane and functionalized with the TNT analog, trinitrobenzene sulfonic acid (TNBSA). A Cy5-labeled anti-TNT antibody was incubated within the trinitrobenzene (TNB) modified microcapillary until binding equilibrium, forming the TNB-anti-TNT antibody complex. The TNB-microcapillary was then subjected to a constant flow of buffer as samples were applied. As samples containing TNT were introduced into the microcapillary flow stream, the Cy5-labeled antibody bound to the immobilized TNB was displaced and fluorescence measured by a fluorometer. Using the microcapillary-based reversed-displacement immunosensor (MRDI) format, parts-per-trillion (pptr) detection limits for TNT in seawater (0.25ng/mL or 250pptr) was achieved with an analysis time under 5min. Reported also are experimental results comparing silane deposition conditions and the effect on immunoassay performance and the comparison of fluorescence signal response with the MRDI using four different monoclonal anti-TNT antibodies.