Monoclonal antibody-based ELISAs for part-per-billion determination of polycyclic aromatic hydrocarbons: effects of haptens and formats on sensitivity and specificity.

As a first step toward developing sensitive enzyme-linked immunosorbent assays (ELISAs) for multianalyte detection of polycyclic aromatic hydrocarbons (PAHs), haptens with different lengths of carboxylic acid spacers at various positions were derived from naphthalene, fluorene, anthracene, phenanthrene, pyrene, fluoranthene, chrysene, and benzo[a]pyrene (BaP). These haptens were coupled with bovine serum albumin (BSA) to form competitor conjugates. All of these haptens were recognized to different extents by monoclonal antibodies (MAbs) 4D5 and 10C10 originally derived by Gomes and Santella (Chem. Res. Toxicol. 1990, 3, 307-310). The most sensitive indirect ELISAs were obtained by coating wells with the least competitive conjugates. Direct ELISAs using horseradish peroxidase conjugates of pyrene and BaP were less sensitive. The MAbs bound BaP with spacers at either C1 or C6. The cross-reactivity profiles of the eight PAHs were different with each PAH-BSA conjugate used as coating antigen. The ELISA results for BaP closely correlated with those by gas chromatography (GC), but the detection limit of the ELISA was approximately 150-fold more sensitive than that of GC, with 2-600 nM spike recoveries of 80-127% from human urine and canal and tap water.