A sandwich ELISA for measuring benzo[a]pyrene–albumin adducts in human plasma

Exposure to polycyclic aromatic hydrocarbons has often been quantified via DNA or human serum albumin (HSA) adducts of the carcinogenic metabolite benzo[a]pyrene diol epoxide (BPDE). We previously reported a sandwich ELISA, using 8E11 as capture antibody and anti-HSA as detection antibody, that detected intact BPDE adducts in HSA isolated from plasma. After confirming that BPDE binds to HSA at His146 and Lys195, we modified the ELISA to measure intact BPDE–HSA directly in human plasma. To adjust for interference due to nonspecifically bound HSA on well surfaces and to cross-reactivity of the antibodies, the ELISA employs paired wells with and without addition of BPDE tetrols to deactivate 8E11. By performing assays in quadruplicate, a series of sample-specific adjustments and screening steps are used to reduce measurement errors that are a consequence of detecting low BPDE–HSA concentrations in the general population. ELISA measurements of BPDE–HSA in plasma from smoking and nonsmoking subjects (range 0.280–2.88 ng BPDE–HSA/mg HSA) and from highway workers with and without exposure to asphalt emissions (range 0.346–13.9 ng BPDE–HSA/mg HSA) detected differences in BPDE–HSA levels in the a priori expected directions.