Development of new rat monoclonal antibodies with different selectivities and sensitivities for 2,4,6-trinitrotoluene (TNT) and other nitroaromatic compounds |
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Authors: | Email author" target="_blank">Petra?M?Kr?merEmail author Elisabeth?Kremmer Cristina?M?Weber Ioan?M?Ciumasu Stephan?Forster Antonius?A?Kettrup |
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Institution: | (1) GSF—National Research Centre for Environment and Health, Institute of Ecological Chemistry, Ingolstädter Landstraße 1, 85764 Neuherberg (Oberschleißheim), Germany;(2) Wissenschaftszentrum Weihenstephan, Lehrstuhl für Ökologische Chemie und Umweltanalytik, Technische Universität München, Weihenstephaner Steig 23, 85350 Freising, Germany;(3) GSF—National Research Centre for Environment and Health, Institute of Molecular Immunology, Marchioninistraße 25, 81377 Munich, Germany |
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Abstract: | Five new rat monoclonal antibodies (mAbs) for 2,4,6-trinitrotoluene (TNT) and other nitroaromatic compounds, including, especially, the metabolite 2-amino-4,6-dinitrotoluene (2-ADNT), are described. Five heterogeneous, competitive enzyme-linked immunosorbent assays (ELISAs) were developed. Assay 1 uses mAb DNT4 3F6 as recognition element and gives a standard curve for TNT in 40 mmol L–1 phosphate buffered saline (PBS) with a test midpoint (IC50) of 0.26±0.08 g L–1 (n=20). Assay 2 (mAb DNT4 4G4) has an IC50 of 0.35±0.07 g L–1 (n=18), assay 3 (mAb DNT4 1A3) has an IC50 of 0.73±0.14 g L–1 (n=15), and assay 4 (mAb DNT4 1A7) has an IC50 of 2.32±0.70 g L–1 (n=15). Assay 5 (mAb DNT2 4B4) is very selective for 2-ADNT and has an IC50 of 8.5±1.7 g L–1 (n=15) in PBS. These antibodies for nitroaromatic compounds differ not only in their sensitivity but also in their selectivity. Major cross-reactants are 1,3,5-trinitrobenzene, 2-ADNT, 4-amino-2,6-dinitrotoluene (4-ADNT), 2,4-dinitroaniline, 3,5-dinitroaniline, and 2,6-dinitroaniline. Although assay 5 is not highly sensitive, the mAb DNT2 4B4 in this assay is highly selective for 2-ADNT. Of all the compounds tested, only 2,4-dinitroaniline and 3,5-dinitroaniline had relevant cross reactivities, 18% and about 26%, respectively. Two ELISAs, using mAbs DNT4 3F6 and DNT2 4B4, were used to analyze different concentrations of TNT and 2-ADNT, respectively, in three different surface water matrices (river and lake water). Both assays were affected by the matrix, but usually performed well (recovery within the range 70–120%). In addition, these ELISAs were used to analyze mixtures of TNT, 2-ADNT, and 4-ADNT, at three different concentrations, in the same water matrices. A different recognition pattern was clearly visible with both assays and depended on the cross reactivities of the corresponding mAb.Dedicated to the memory of Wilhelm Fresenius |
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Keywords: | TNT 2-Amino-4 6-dinitrotoluene 4-Amino-2 6-dinitrotoluene Nitroaromatic compounds Immunoassay Monoclonal antibodies Water Pattern recognition |
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