Electrochemiluminescence enzyme immunoassays for TNT and pentaerythritol tetranitrate

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.