Development of new rat monoclonal antibodies with different selectivities and sensitivities for 2,4,6-trinitrotoluene (TNT) and other nitroaromatic compounds.
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 microg L(-1) (n=20). Assay 2 (mAb DNT4 4G4) has an IC50 of 0.35+/-0.07 microg L(-1) (n=18), assay 3 (mAb DNT4 1A3) has an IC50 of 0.73+/-0.14 microg L(-1) (n=15), and assay 4 (mAb DNT4 1A7) has an IC50 of 2.32+/-0.70 microg L(-1) (n=15). Assay 5 (mAb DNT2 4B4) is very selective for 2-ADNT and has an IC50 of 8.5+/-1.7 microg 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.
Krämer
Petra M
PM
GSF, National Research Centre for Environment and Health, Institute of Ecological Chemistry, Ingolstädter Landstrasse 1, 85764 Neuherberg, Oberschleissheim, Germany. kramer@gsf.de
Kremmer
Elisabeth
E
Weber
Cristina M
CM
Ciumasu
Ioan M
IM
Forster
Stephan
S
Kettrup
Antonius A
AA
D000814:Aniline Compounds / Q000032:analysis*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D000918:Antibody Specificity; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006241:Haptens / Q000737:chemistry; D009574:Nitro Compounds / Q000032:analysis; D051381:Rats; D012680:Sensitivity and Specificity; D014303:Trinitrotoluene / Q000032:analysis*; D014867:Water / Q000032:analysis
D000814:Aniline Compounds; D000911:Antibodies, Monoclonal; D006241:Haptens; D009574:Nitro Compounds; D014867:Water; D014303:Trinitrotoluene; C035208:2-amino-4,6-dinitrotoluene