A rapid technique for estimating DNA binding, used to evaluate 1-nitropyrene adduct formation.
A simple and rapid procedure for estimating binding of radio-labelled material to DNA and protein is described. Protein was extracted from lysed rabbit alveolar macrophages with chloroform: iso-amyl-alcohol:phenol extraction. Nucleic acids were precipitated from the lysate, and hydrolysed with protease and NaOH to remove residual protein and RNA respectively. Bound radioactivity was quantitated by precipitation of DNA onto glass fiber filters. Protein labelled with 3H-leucine and DNA and RNA adducts formed from 1-nitro[14C]pyrene by xanthine oxidase were used to define this procedure. 14C was shown to be bound to endogeneous protein and DNA isolated from rabbit alveolar macrophages that had been incubated with 1-nitro[14C]pyrene.
D000818:Animals; D002250:Carbon Radioisotopes; D004247:DNA / Q000378:metabolism*; D008264:Macrophages / Q000378:metabolism; D008722:Methods; D011650:Pulmonary Alveoli; D011721:Pyrenes / Q000378:metabolism*; D011817:Rabbits
D002250:Carbon Radioisotopes; D011721:Pyrenes; D004247:DNA; C032668:1-nitropyrene