Jackson M A, King L C, Ball L M
Drug Chem Toxicol. 1983;6(6):549-62. doi: 10.3109/01480548309017809.
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.
本文描述了一种简单快速的方法，用于估算放射性标记物质与DNA和蛋白质的结合。采用氯仿：异戊醇：苯酚萃取法从裂解的兔肺泡巨噬细胞中提取蛋白质。从裂解物中沉淀出核酸，并用蛋白酶和氢氧化钠水解，分别去除残留的蛋白质和RNA。通过将DNA沉淀到玻璃纤维滤膜上来定量结合的放射性。用3H-亮氨酸标记的蛋白质以及由黄嘌呤氧化酶作用于1-硝基[14C]芘形成的DNA和RNA加合物来验证该方法。结果表明，14C与从用1-硝基[14C]芘孵育过的兔肺泡巨噬细胞中分离出的内源性蛋白质和DNA相结合。
