A New Method for the Detection and Measurement of Polyaromatic Carcinogens and Related Compounds by DNA Intercalation

Abstract

A simple and sensitive method for detection and quantification of carcinogens and related polyaromatic compounds by DNA intercalation has been devised. The experimental technique is based on the phenomenon of DNA intercalation using fluorescence polarization for quantitative measurements. The assay architecture is analogous to a protocol presently used for competitive immunoassays, whereby an intercalating dye or fluorochrome would compete with or be displaced by the analyte (test compound). Our competitive binding assay utilizes DNA binding sites and a fluorescent intercalating dye. The carcinogenic analytes investigated were benzo[a]pyrene, dibenz[a,h]anthracene, dibenz[a,j]anthracene, benz[a]anthracene, benzo[j]fluoranthene, benzidine, aniline, parathion, pentachlorophenol and nitrobenzene. The noncarcinogenic hydrocarbons studied were naphthalene, anthracene, phenanthrene, benzo[k]fluoranthene and 1,2,3,4,5,6,7,8-octahydronaphthalene. The intercalating dyes examined were acridine orange, ethidium bromide, proflavin and 4,6-diamidino 2-phenylindale chloride (DAPI). Of the compounds studied, only the single ring compounds gave a negative response. All the other compounds investigated displaced the intercalated dye molecule. It has been determined that molecules require at least two adjacent benzene rings for intercalation to occur. The detection limits of this analytical method were between 10-5 mol/L and 10-8 mol/L for all materials tested.