Electrochemiluminescence Enzyme Immunoassays for TNT and Pentaerythritol Tetranitrate

Department of Chemistry, Liverpool University, Liverpool L69 7ZD, UK.
Analytical Chemistry (Impact Factor: 5.64). 09/2003; 75(16):4244-9. DOI: 10.1021/ac034163s
Source: PubMed


Electrochemiluminescence enzyme immunoassays for 2,4,6-trinitrotoluene (TNT) and pentaerythritol tetranitrate (PETN) are described. The latter is, to the best of our knowledge, the first report of an immunoassay for PETN. Haptens corresponding to these explosives were covalently attached to high-affinity dextran-coated paramagnetic beads. The beads were mixed with the corresponding Fab fragments and the sample. After adding a second HRP-labeled antispecies-specific antibody, the mixture was pumped into an electrochemiluminometer where beads were concentrated on the working electrode magnetically. The amount of analyte in the sample was determined by measuring light emission when H2O2 was generated electrochemically in the presence of luminol and an enhancer. The detection limits for TNT and PETN were 0.11 and 19.8 ppb, respectively. Details of bead preparation and performance are given. The increase in sensitivity obtained when Fab fragments are used instead of whole antibodies is explained, and the implications of this observation for nanoparticle-based assays are discussed.

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    • "The antibody was briefly mentioned in a paper by Blackburn et al. (Blackburn et al., 2000); however, no details were given. The monoclonal antibodies were used by Wilson et al. (Wilson et al., 2003) for an electrochemiluminescence enzyme immunoassay reaching a detection limit of about 20 μg/l and some other work (Aojula et al., 2002). The hapten design for PETN antibodies is particularly difficult, because the molecule shows high symmetry and does not offer any site for chemical modification. "
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    ABSTRACT: An improved antibody against the explosive pentaerythritol tetranitrate (PETN) was developed. The immunogen was designed by the concept of bioisosteric replacement, which led to an excellent polyclonal antibody with extreme selectivity and immunoassays of very good sensitivity. Compounds such as nitroglycerine, 2,4,6-trinitrotoluene, 1,3,5-trinitrobenzene, hexogen (RDX), 2,4,6-trinitroaniline, 1,3-dinitrobenzene, octogen (HMX), triacetone triperoxide, ammonium nitrate, 2,4,6-trinitrophenol and nitrobenzene were tested for potential cross-reactivity. The detection limit of a competitive enzyme-linked immunosorbent assay was determined to be around 0.5 µg/l. The dynamic range of the assay was found to be between 1 and 1000 µg/l, covering a concentration range of three decades. This work shows the successful application of the bioisosteric concept in immunochemistry by exchange of a nitroester to a carbonate diester. The antiserum might be used for the development of quick tests, biosensors, microtitration plate immunoassays, microarrays and other analytical methods for the highly sensitive detection of PETN, an explosive frequently used by terrorists, exploiting the extreme difficulty of its detection. Copyright © 2015 John Wiley & Sons, Ltd.
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    • "Fully Electrical Microarrays 267 explosives, biological toxins and antibiotics is the main goal (Pestka, 1991; Beier and Stanker, 2000; Loomans et al., 2003; Wilson et al., 2003; Estevez–Alberola and Marco, 2004; Lee et al., 2004). To achieve high sensitivities, physical test methods like high-pressure liquid chromatography (HPLC) or gas chromatography with mass detection (GC–MS) are enabled (Sorensen et al., 2003). "

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