A SENSITIVE COLOR ELISA FOR DETECTING POLYCYCLIC AROMATIC HYDROCARBON-DNA ADDUCTS IN HUMAN TISSUES

Human exposure to polycyclic aromatic hydrocarbons (PAHs) has been det ermined by measurement of DNA adducts in human tissues, Competitive en zyme-linked immunosorbent assays (ELISAs) using antisera recognizing b enzo[a]pyrenediol-epoxide-modified DNA (BPDE-I-DNA) and color or fluor escence endpoint detection have been used extensively for quantifying PAH-DNA adducts, The fluorescence ELISA (limit of detection 1 adduct/1 0(8) nucleotides) was previously reported to be more sensitive than th e color ELISA (1/10(7)) for measuring PAH adducts (Santella et al. (19 88) Carcinogenesis, 9, 1265-1269). However, the fluorescence assay has the disadvantages of greater variation among the replicates and highe r background levels than the color assay. Using a newly developed anti serum against BPDE-I-DNA, we have modified the color ELISA so that it has the same sensitivity as the fluorescence ELISA and requires only 3 3% of the sample quantity needed for the fluorescence ELISA. The modif ications included preincubation of the antiserum with the samples, usi ng microtiter plates with half-size, flat bottom wells, and optimizing the assay conditions, The improved color ELISA was used to analyze DN A samples from human autopsy tissues, including heart, lung, Liver, ki dney, spleen, pancreas and stomach from smokers and nonsmokers. With t he exception of spleen and stomach, all tissues from smokers showed hi gher PAH-DNA adducts (ranging from 0.3 to 19.0 adducts/10(7) nucleotid es) than the tissues from the nonsmokers (0.3 to 3.7 adducts/10(7) nuc leotides) in two separate experiments, Among the tissues from smokers, heart showed the highest level of DNA adducts. This study demonstrate s that a stable color ELISA with high sensitivity can be useful in ass essing human exposure to PAH.