Development of an Enzyme-Linked Immunosorbent Assay for 1-Nitropyrene: A Possible Marker Compound for Diesel Exhaust Emissions

An enzyme-linked immunosorbent assay (ELISA) was developed for 1-nitropyrene. For hapten syntheses pyrenebutyric acid was nitrated yielding a mixture of 4-(6-nitropyrene-1-yl)- and 4-(8-nitropyrene-1yl)-butyric acid isomers that could not be separated. High titered antisera were obtained in rabbits after immunization with a KLH immunogen prepared with the mixture of isomers. As coating antigen a corresponding BSA conjugate was used. The calibration curve was linear in the 0.1 to 10 ppb range with a center point at 1.5 ppb and an LOD of 0.02 ppb in 10:90 acetonitrile/water (v/v). The average coefficients of variation were 4.9 and 7.8% over the range of the standard curve for intra- and interassay precision. Interference caused by 1,6- and 1,8-dinitropyrenes was observed. The effect of different concentrations of solvents on the antibody-analyte interaction was studied.